| replication of orgyia pseudotsugata baculovirus dna: lef-2 and ie-1 are essential and ie-2, p34, and op-iap are stimulatory genes. | a transient dna replication assay was used to identify genes located within m.u. 90.5-7.0 of the orgyia pseudotsugata multinucleocapsid nuclear polyhedrosis virus (opmnpv) genome that influenced replication of a reporter plasmid containing an opmnpv origin of replication, when cotransfected into uninfected lymantria dispar cells. the viral transactivator ie-1 and a 2.4-kb subclone were found to be essential for replication. the 2.4-kb region was sequenced and open reading frames were identified. ... | 1995 | 7571435 |
| evidence for proteolytic processing of tobacco mosaic virus movement protein in arabidopsis thaliana. | two ecotypes of arabidopsis thaliana were transformed with the gene encoding tobacco mosaic virus (tmv) movement protein (p30). p30 accumulated largely in a subcellular fraction containing cell wall components and as a soluble protein. the protein migrated in denaturing gels with an m(r) of 30k, significantly faster than p30 (m(r) approximately 34k) accumulating after expression in transgenic tobacco, escherichia coli or spodoptera frugiperda cells, or after virus multiplication in tobacco. the ... | 1995 | 7579611 |
| cdna-directed expression of human cytochrome p450 cyp1a1 using baculovirus. purification, dependency on nadph-p450 oxidoreductase, and reconstitution of catalytic properties without purification. | a recombinant baculovirus containing the human cytochrome p450 (cyp) 1a1 cdna was constructed and used to express cyp1a1 in spodoptera frugiperda (sf9) insect cells (0.14 +/- 0.04 nmol/mg protein, 53 +/- 14 nmol/liter, n = 30). the enzyme represented approximately 1% of total cellular protein and was partially purified by a three-column procedure to a specific content of 5.0 nmol/mg protein. catalytic activity was reconstituted with both the purified enzyme using lipid and nadph-p450 oxidoreduct ... | 1995 | 7587956 |
| [direct expression of the pig testicular 3 alpha/beta(20 beta)- hydroxysteroid dehydrogenase cdna using baculovirus expression system]. | the direct expression of the pig testicular 3 alpha/beta(20 beta)- hydroxysteroid dehydrogenase (hsd) cdna using a baculovirus expression system was investigated. a minimum essential cdna containing the coding region of 3 alpha/beta(20 beta)-hsd was amplified by polymerase chain reaction (pcr) to extend the dna linker including the bam hi restriction site in the both ends of the cdna. as the template, 3 alpha/beta(20 beta)-hsd cdna, pbs 52, which was subcloned to bluescript ii was used. the pcr ... | 1995 | 7602499 |
| expression and characterization of the zeta 1 subunit of the n-methyl-d-aspartate (nmda) receptor channel in a baculovirus system. | using a baculovirus expression vector system, the zeta 1 subunit of the mouse n-methyl-d-aspartate (nmda) receptor channel was expressed in spodoptera frugiperda insect cells. the peptide corresponding to the c-terminus of the zeta 1 subunit was synthesized by using the multiple antigen peptide (map) system, and an antibody to the synthetic peptide was produced. immunoblotting using the newly developed antibody revealed the major 122-kda and the minor 104-kda protein bands. the effect of tunicam ... | 1995 | 7609635 |
| production and purification of recombinant african swine fever virus attachment protein p12. | the conditions for cultivation of spodoptera frugiperda (sf9) insect cells for production of recombinant baculoviruses have been studied, to scale-up and improve the efficiency of the process for production of the african swine fever virus attachment protein p12 in the baculovirus expression system. it was shown that the total virus and recombinant protein production in insect cells infected with the acp12 recombinant baculovirus were slightly dependent on cell density, but largely dependent on ... | 1995 | 7612242 |
| molecular structure and function of cd4 on murine egg plasma membrane. | in the present study, the expression of the cd4 molecule on murine egg plasma membrane was confirmed by the indirect immunofluorescence (iif) method. the full-length cd4 cdna from murine eggs was synthesised by the reverse transcriptase-polymerase chain reaction (rt-pcr) method and its authenticity verified by southern blot hybridisation using an end-labelled internal oligonucleotide. the results of dna sequencing showed that the nucleotide sequence of the cdna of cd4 from murine egg mrna was id ... | 1995 | 7613876 |
| ligand binding properties of muscarinic acetylcholine receptor subtypes (m1-m5) expressed in baculovirus-infected insect cells. | five subtypes of muscarinic acetylcholine receptors (m1-m5) have been expressed in insect cells (spodoptera frugiperda, sf9) using the baculovirus system. up to 6 nmol of muscarinic acetylcholine receptors were produced by 1 liter culture; 0.3 to 0.6 (human m1), 3 to 6 (human m2), 2 to 4 (rat m3), 1 to 2 (rat m4) and 0.5 to 1 (human m5) nmol. pirenzepine, af-dx116 and hexahidrosiladifenidol showed the highest affinity for the m1, m2 and m3 subtype, respectively, indicating that these receptors e ... | 1995 | 7616422 |
| nucleotide labeling and reconstitution of the recombinant 58-kda subunit of the vacuolar proton-translocating atpase. | evidence suggests that atp hydrolysis catalyzed by the clathrin-coated vesicle proton-translocating atpase requires at least four polypeptides of molecular masses of 70, 58, 40, and 33 kda (xie, x.-s., and stone, d.k. (1988) j. biol. chem. 263, 9859-9867). to further investigate the subunit requirements for atp hydrolysis, histidine-tagged, 58-kda polypeptide was expressed in insect sf9 (spodoptera frugiperda) cells. after purification by ni(2+)-nitrolotriacetic acid chromatography, the 58-kda p ... | 1995 | 7622510 |
| heterologous expression and enzymatic properties of a selenium-independent glutathione peroxidase from the parasitic nematode brugia pahangi. | a full-length cdna from the parasitic nematode brugia pahangi encoding a secreted homolog of glutathione peroxidase in which the codon for the active site selenocysteine is substituted naturally by a cysteine codon has been expressed in spodoptera frugiperda (insect) cells via autographa californica nuclear polyhedrosis virus (baculovirus). the recombinant protein was glycosylated and secreted from the cells in tetrameric form. the purified protein showed glutathione peroxidase activity with a r ... | 1995 | 7629152 |
| horseradish peroxidase phe172-->tyr mutant. sequential formation of compound i with a porphyrin radical cation and a protein radical. | a gene coding for the f172y mutant of horseradish peroxidase isozyme c (hrp) has been constructed and expressed in both spodoptera frugiperda (sf-9) and trichoplusia ni egg cell homogenate (highfive) cells. homology modeling with respect to three peroxidases for which crystal structures are available places phe172 on the proximal side of the heme in the vicinity of porphyrin pyrrole ring c. the ph optimum and spectroscopic properties of the f172y mutant are essentially identical to those of wild ... | 1995 | 7629167 |
| expression of the human 5-hydroxytryptamine1a receptor in sf9 cells. reconstitution of a coupled phenotype by co-expression of mammalian g protein subunits. | the possibility that spodoptera frugiperda (sf9) cells can provide an intact cell setting for reconstitution of the human 5-hydroxytryptamine1a (5-ht1a) receptor with mammalian g protein subunits was explored. the 5-ht1a receptor was found to assume an uncoupled phenotype when expressed alone in sf9 cells at relatively high levels (5-34 pmol of receptor/mg of membrane protein), i.e. agonist-binding to the receptor was characterized by a relatively high kd and an insensitivity to gtp. co-expressi ... | 1995 | 7629202 |
| construction and adhesive properties of a soluble madcam-1-fc chimera expressed in a baculovirus system: phylogenetic conservation of receptor-ligand interaction. | madcam-1 is a high endothelial venule adhesion molecule composed of immunoglobulin and mucin-like domains which binds the leucocyte integrin lpam-1 (alpha 4 beta 7), and is largely responsible for the selective homing of lymphocytes to mucosal tissues. a novel soluble form of mouse madcam-1 which is normally membrane bound has been produced by joining the extracellular region of the receptor to the fc domain of human igg1. the madcam-1-fc cdna was inserted into the genome of autographa californi ... | 1995 | 7631157 |
| comparison of the enzymatic properties of the na,k-atpase alpha 3 beta 1 and alpha 3 beta 2 isozymes. | the coexpression of multiple isoforms of the alpha and beta subunits of the na,k-atpase in mammalian tissues gives rise to the complex molecular heterogeneity that characterizes the na pump. the expression of the different na,k-atpase isoforms in insect cells using recombinant baculoviruses represents a useful system for the analysis of na,k-atpase isoform function. in the present study, we use this system to direct the expression of the rat na,k-atpase alpha 3 beta 1 and alpha 3 beta 2 in sf-9 ... | 1995 | 7632689 |
| baculovirus-driven expression and purification of glycine receptor alpha 1 homo-oligomers. | the glycine receptor is a ligand-gated anion channel protein of postsynaptic membranes. we expressed a homo-oligomeric receptor composed of human alpha 1 subunits in spodoptera frugiperda cells by infection with a recombinant autographa californica nuclear polyhedrosis virus. a substantial fraction of the recombinant receptor was incorporated as a functional channel protein into the cell's plasma membrane at expression levels 4- to 30-fold higher than in other eukaryotic heterologous expression ... | 1995 | 7635206 |
| expression of pe38 and ie2, viral members of the c3hc4 finger family, during baculovirus infection: pe38 and ie2 localize to distinct nuclear regions. | the pe38 gene of autographa californica nuclear polyhedrosis virus represents one of the major early transcripts after viral infection. the function of the pe38 protein, which contains a c3hc4 zinc finger motif, is still not understood. we have raised polyclonal antiserum against the pe38 protein, pe38, produced in bacteria to investigate pe38 expression in the course of infection. a approximately 38-kda polypeptide is first detectable at 2 h postinfection and decreases rapidly after 24 h. durin ... | 1995 | 7636971 |
| the rotavirus nonstructural glycoprotein nsp4 mobilizes ca2+ from the endoplasmic reticulum. | we previously reported that expression of rotavirus nonstructural glycoprotein nsp4 is responsible for an increase in cytosolic free ca2+ concentration ([ca2+]i) in spodoptera frugiperda (sf9) insect cells (p. tian, y. hu, w. p. schilling, d. a. lindsay, j. eiden, and m. k. estes, j. virol. 68:251-257, 1994). the purpose of the present study was to determine the mechanism by which nsp4 causes an increase in [ca2+]i by measuring the permeability of the cytoplasmic and endoplasmic reticulum (er) m ... | 1995 | 7637021 |
| isolation and characterization of c-fos recombinant baculovirus. | the use of the baculovirus system to produce recombinant proteins is based on the high level of protein production and the possibility to obtain, in spodoptera frugiperda insect cells, recombinant proteins with the post-translational modifications found in the native proteins. here we describe the isolation and characterization of a recombinant baculovirus containing the mouse c-fos gene. the c-fos cdna was subcloned into the pvl1392 baculovirus transfer vector. the recombinant plasmid (pvl1392. ... | 1994 | 7640625 |
| characterization of two human camp-specific phosphodiesterase subtypes expressed in baculovirus-infected insect cells. | recombinant baculoviruses were constructed to express cdnas encoding two distinct subtypes of human camp-specific phosphodiesterase (hpde4a and hpde4b). infection of spodoptera frugiperda insect cells with the appropriate recombinant baculoviruses resulted in high level production of biologically-active protein as measured by enzymatic activity and immunoblotting using subtype-specific anti-hpde4 antisera. both recombinant proteins showed catalytic activity with a low km (approximately 3 microm) ... | 1995 | 7640661 |
| comparison of the non-structural protein, ns1, of tick-borne and insect-borne orbiviruses. | the nucleotide sequence of rna segment 6 of broadhaven virus (brdv), a tick-borne orbivirus, was determined principally from two overlapping cdna clones and rna end sequence analysis. the genome segment is 1714 base pairs in length and has a coding capacity for a protein of 537 amino acids, having a net charge of +4.0 at neutral ph. comparison of the predicted amino acid sequence of brdv rna segment 6 with the ns1 sequence of insect-borne orbiviruses, bluetongue virus (btv), african horse sickne ... | 1995 | 7653106 |
| association of a 33-kilodalton cysteine proteinase found in corn callus with the inhibition of fall armyworm larval growth. | protein patterns of callus from corn (zea mays l.) inbreds that are either resistant or susceptible to fall armyworm (spodoptera frugiperda [j.e. smith]) were analyzed by two-dimensional electrophoresis. fall armyworm larvae reared on callus initiated from resistant inbreds were significantly smaller than those reared on callus of susceptible inbreds. a 33-kd protein found in callus from the resistant inbreds mp704 and mp708 was absent in callus from the susceptible inbreds tx601 and ab24e. howe ... | 1995 | 7659755 |
| efficient complementary dna directed expression of human fetal liver cytochrome p450 (cyp3a7) in insect cells using baculovirus. | cyp3a7 is a form of cytochrome p450, which is expressed specifically in human fetal livers. npvhf1, a recombinant baculovirus containing the entire coding region of cyp3a7, was constructed and infected to spodoptera frugiperda (sf9) cells. upon infection with npvhf1, the sf9 cells expressed the cyp3a7 to the maximum content of 0.2 nmol per mg of whole cell lysates 72 hours after infection. a 5.5-fold expression level (1.1 nmol per mg of whole cell lysates) was attainable when cultured in the pre ... | 1995 | 7663401 |
| differential requirements for baculovirus late expression factor genes in two cell lines. | a plasmid library of 18 late expression factor (lef) genes (lef library) from the baculovirus autographa californica nuclear polyhedrosis virus (acmnpv) supports transient expression from a late viral promoter in the sf-21 cell line, derived from spodoptera frugiperda. we found, however, that this lef library was unable to support expression from the same promoter in the tn-368 cell line, derived from trichoplusia ni, which is also permissive for acmnpv replication. to identify the additional fa ... | 1995 | 7666527 |
| high-level secretion of native rat pancreatic lithostathine in a baculovirus expression system. | we have constructed a recombinant baculovirus expression vector containing rat pancreatic lithostatin cdna. baculovirus infection of spodoptera frugiperda (sf9) insect cells resulted in the de novo synthesis and secretion of a recombinant protein demonstrating an apparent molecular weight of about 16.5 kda. under optimal conditions [multiplicity of infection of 5 plaque-forming units (pfu)/cell and culture times of 48-56 h postinfection] recombinant protein was secreted into the culture medium a ... | 1995 | 7667245 |
| morphological changes of the insect cells in the baculovirus system as a function of v-myb and c-myb inserts expression and topographic localization of v-myb and c-myb proteins. | structural changes of insect cells spodoptera frugiperda in the baculovirus expression system after expression of v-myb oncogene and c-myb protooncogene inserts were studied by means of electron microscopy. expression of v-myb gene insert was accompanied by extensive changes in cell structure, when compared with those of the noninfected and wild-type virus-infected cells. enormous increase in nuclear content was apparent within 48 hrs after infection, along with changes in nucleolus appearance. ... | 1995 | 7676943 |
| lepidopteran-specific crystal toxins from bacillus thuringiensis form cation- and anion-selective channels in planar lipid bilayers. | previous studies in our laboratory have shown that cryic, a lepidopteran-specific toxin from bacillus thuringiensis, triggers calcium and chloride channel activity in sf-9 cells (spodoptera frugiperda, fall armyworm). chloride currents were also observed in sf-9 membrane patches upon addition of cryic toxin to the cytoplasmic side of the membrane. in the present study the ability of activated cryic toxin to form channels was investigated in a receptor-free, artificial phospholipid membrane syste ... | 1993 | 7681482 |
| high level expression and phosphorylation of hepatitis b virus polymerase in insect cells with recombinant baculoviruses. | the hepatitis b virus polymerase open reading frame, as well as various subdomains of polymerase, was expressed in insect cells using the recombinant baculovirus expression system. full-length polymerase was expressed at very low levels in a spodoptera frugiperda cell line, the amino-terminal domain of polymerase was expressed at high levels, and other constructs were expressed at intermediate levels. infections of a trichoplusia ni cell line with the same recombinant baculoviruses resulted in h ... | 1993 | 7683161 |
| expression of a recombinant baculovirus for vitronectin in insect cells: purification, characterization of post-translational modifications and functional studies of the recombinant protein. | vitronectin (vn) is a cell attachment glycoprotein present in plasma and the extracellular matrix that also has multiple regulatory roles in the complement, coagulation, and fibrinolytic systems. vn stabilizes plasminogen activator inhibitor-1 (pai-1) with potential binding sites reported at both the somatomedin b and heparin binding domains. in order to perform detailed structure-function analyses of vn, we have produced and expressed a recombinant baculovirus containing the cdna of human vn. r ... | 1993 | 7688496 |
| expression of the glycosylphosphatidylinositol-linked complement-inhibiting protein cd59 antigen in insect cells using a baculovirus vector. | cd59 antigen (cd59) is a glycosylphosphatidylinositol (gpi)-linked membrane glycoprotein which protects human cells from complement-mediated lysis. here we report the expression of functionally active cd59 in spodoptera frugiperda insect cells using a baculovirus vector. recombinant cd59 was expressed abundantly on the surface of the insect cells and protected the cells from lysis by human complement. the protein was released from the cell surface by treatment with phosphatidylinositol-specific ... | 1993 | 7694573 |
| ion channel activity of n-terminal fragments from cryia(c) delta-endotoxin. | proteolytically-derived amino-terminal fragments from the bacillus thuringiensis delta-endotoxin, cryia(c), demonstrated ion channel activity in two in vitro assays in the absence of any membrane receptors. a 24.5 kda fragment (from spodoptera frugiperda digestive proteolysis) formed cation-selective channels (10mv for a 3:1 salt gradient) in planar lipid bilayers up to several hundred picosiemen conductance. a 21.5 kda fragment doublet (from alkaline hydrolysis) also showed large conductance in ... | 1993 | 7694582 |
| expression of the newcastle disease virus (ndv) fusion glycoprotein and vaccination against ndv challenge with a recombinant baculovirus. | the hybrid baculovirus constructed from autographa california nuclear polyhedrosis virus (npv) and bombyx mori (silkworm) npv was used for expression of fusion glycoprotein (f) of newcastle disease virus (ndv) strain d26. the gene encoding f protein was introduced into the improved baculovirus expression vector derived from the host-range-expanded baculovirus. in spodoptera frugiperda (fall armyworm) cells infected with a recombinant baculovirus, hyf121, the expressed f protein was properly loca ... | 1994 | 7702510 |
| overexpression, purification, and characterization of full-length and mutant caldesmons using a baculovirus expression system. | three recombinant chicken gizzard caldesmon (cad) baculovirus vectors that contained the full-length cad codon sequence (pv1cad), the full-length cad codon sequence and a six-histidine tag at the 5'-end (pbluebachiscad), or the full-length cad codon sequence and an extra six-histidine codon sequence at the 3'-end (pvlhiscad) were constructed. spodoptera frugiperda (sf9) cells transfected with these constructs overexpressed full-length cad, yielding 2, 20, and 50 micrograms per 10(6) cells for pb ... | 1994 | 7706421 |
| differential transcription of baculovirus late and very late promoters: fractionation of nuclear extracts by phosphocellulose chromatography. | an in vitro transcription system based on cytidine-free cassette was developed for the late 39k gene and the very late polyhedrin gene of autographa californica nuclear polyhedrosis virus (acnpv). optimization of transcription conditions revealed that a preincubation step was not required for transcription of late and very late promoters, although preincubation was essential for efficient transcription from an early promoter. the 39k and polyhedrin constructs were actively transcribed by nuclear ... | 1995 | 7707516 |
| molecular and biochemical evidence for the involvement of the asp-333-his-523 pair in the catalytic mechanism of soluble epoxide hydrolase. | in order to investigate the involvement of amino acids in the catalytic mechanism of the soluble epoxide hydrolase, different mutants of the murine enzyme were produced using the baculovirus expression system. our results are consistent with the involvement of asp-333 and his-523 in a catalytic mechanism similar to that of other alpha/beta hydrolase fold enzymes. mutation of his-263 to asparagine led to the loss of approximately half the specific activity compared to wild-type enzyme. when his-3 ... | 1995 | 7713895 |
| molecular cloning and expression of catrocollastatin, a snake-venom protein from crotalus atrox (western diamondback rattlesnake) which inhibits platelet adhesion to collagen. | a 50 kda protein that inhibits platelet adhesion to collagen has been isolated from snake venom of crotalus atrox (western diamondback rattlesnake) and has been named 'catrocollastatin'. the cdna cloning of catrocollastatin has been accomplished. a full-length cdna of 2310 bp with an open reading frame between nucleotides 51 and 1880 was obtained. the deduced amino acid sequence consists of 609 amino acids. the cdna-predicted amino acid sequence is highly similar to that of haemorrhagic metallop ... | 1995 | 7733877 |
| overexpression of bombyx mori prothoracicotropic hormone using baculovirus vectors. | recombinant baculoviruses were constructed that express the cdna encoding the prothoracicotropic hormone (ptth) of bombyx mori. this hormone stimulates the production of ecdysteroids by the insect's prothoracic glands. two groups of viruses were constructed, expressing either the entire cdna encoding prepro-ptth, or a synthetic chimeric gene encoding a signal peptide fused to the mature ptth subunit. in both cases, the genes were expressed in wild-type autographa californica nuclear polyhedrosis ... | 1995 | 7742834 |
| functional expression of a diuretic hormone receptor in baculovirus-infected insect cells: evidence suggesting that the n-terminal region of diuretic hormone is associated with receptor activation. | a recombinant baculovirus containing the diuretic hormone receptor cdna from manduca sexta was constructed. when spodoptera frugiperda (sf9) cells were infected with the virus, a time-dependent expression of the receptor appeared. the expressed receptor displayed high affinity for diuretic hormone, which was similar to the affinity observed in malpighian tubules and transfected cos-7 cells. the receptor expression level was 77 pmol/mg protein, as compared to 3.1 pmol/mg protein in malpighian tub ... | 1995 | 7742837 |
| histidine tagging both allows convenient single-step purification of bovine rhodopsin and exerts ionic strength-dependent effects on its photochemistry. | for rapid single-step purification of recombinant rhodopsin, a baculovirus expression vector was constructed containing the bovine opsin coding sequence extended at the 3'-end by a short sequence encoding six histidine residues. recombinant baculovirus-infected spodoptera frugiperda cells produce bovine opsin carrying a c-terminal histidine tag (v-opshis6x). the presence of this tag was confirmed by immunoblot analysis. incubation with 11-cis-retinal produced a photosensitive pigment (v-rhohis6x ... | 1995 | 7744755 |
| the autographa californica nuclear polyhedrosis virus homologous region 1a: identical sequences are essential for dna replication activity and transcriptional enhancer function. | hr1a consists of two 30-bp imperfect palindrome sequences separated by 58 bp and each palindrome contains a naturally occurring ecori site at its core. plasmid subclones of the hr1a-containing acmnpv hindiii-n fragment were examined for their ability to replicate in virus-infected (spodoptera frugiperda) sf9 cells, and to stimulate transcription when linked in cis with a 39k gene promoter-beta-glucuronidase fusion and cotransfected into cells along with a plasmid (ple-1) containing the gene enco ... | 1995 | 7747446 |
| cloning and expression of the nucleoprotein of peste des petits ruminants virus in baculovirus for use in serological diagnosis. | peste des petits ruminants (ppr) is a viral disease of goats and sheep characterized by erosive stomatitis, enteritis, and pneumonia. the virus is a member of the family paramyxoviridae and the genus morbillivirus. the disease has high morbidity and mortality rates and has a substantial economic impact in developing countries. we have cloned and sequenced the cdna of the nucleocapsid (n) gene of the nigeria 75/1 strain of ppr virus (pprv). a comparison of its nucleotide and deduced amino acid se ... | 1995 | 7747450 |
| synthesis and processing of the rubella virus p110 polyprotein precursor in baculovirus-infected spodoptera frugiperda cells. | in order to study the processing of rubella virus (rv) structural proteins (capsid protein, of 33 kda; e2 of 42-47 kda; and e1 of 58 kda) in spodoptera frugiperda (fall armyworm) cells, a 24s cdna encoding the polyprotein precursor, p110, was inserted under the transcriptional regulation of the polyhedrin gene promoter of the autographa californica nuclear polyhedrosis virus (acnpv) and expressed during viral infection. by immunoblot analysis using antibodies directed against whole rv and the in ... | 1995 | 7754676 |
| expression of trypanosoma congolense antigens in spodoptera frugiperda insect cells. | transcripts which encode two metacyclic-form-specific variable surface glycoproteins (mvsgs) of trypanosoma congolense il3000 have been cloned into baculovirus expression vectors using a novel transfer vector, pacl11. one of the recombinant baculoviruses (acvsg1) expressed a mvsg as a glycoprotein with a signal peptide which was cleaved in this expression system, whereas the other one (acvsg2) expressed an unprocessed protein. from 1 liter of culture containing 10(9) spodoptera frugiperda cells ... | 1995 | 7758544 |
| protection of guinea-pigs from experimental rocky mountain spotted fever by immunization with baculovirus-expressed rickettsia rickettsii rompa protein. | baculovirus recombinants that express the rickettsia rickettsii rompa protein were constructed. monoclonal antibodies (mabs) against the rompa protein reacted with recombinant-infected spodoptera frugiperda (sf9) cells in indirect immunofluorescence assays. sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting of infected sf9 cell lysates with a mab against rompa showed that the recombinant-expressed rompa protein migrated slightly below rompa extracted from r. rickettsii. ... | 1995 | 7762273 |
| expression of marek's disease virus (mdv) serotype 2 gene which has partial homology with mdv serotype 1 pp38 gene. | we constructed the recombinant baculovirus expressing the gene of non-pathogenic marek's disease virus (mdv) serotype 2 (mdv2) which encodes a polypeptide with partial homology to mdv serotype 1 (mdv1) pp38, an antigen associated with transformed cells. the recombinant mdv2 protein was detected as a band of 32 kda in immunoblot analysis with mdv2-infected chicken serum. mouse serum against insect spodoptera frugiperda cells infected with the recombinant baculovirus immunoprecipitated a 38 kda mo ... | 1995 | 7762295 |
| effect of temperature and oxygen on cell growth and recombinant protein production in insect cell cultures. | the effect of temperature and o2 saturation on the production of recombinant proteins beta-galactosidase and human glucocerebrosidase by spodoptera frugiperda cells (sf9) infected with recombinant autographa californica nuclear polyhedrosis virus was investigated. the rates of cell growth, glucose consumption, o2 consumption and product expression were measured at temperatures between 22 degrees c and 35 degrees c. the results indicated that possible o2 limitation may be alleviated without compr ... | 1993 | 7763472 |
| insect cell hosts for baculovirus expression vectors contain endogenous exoglycosidase activity. | four different insect cell lines that can be used as hosts for baculovirus infection were assayed for the presence of endogenous exoglycosidases. all four cell lines, derived from spodoptera frugiperda, trichoplusia ni, bombyx mori, or malacosoma disstria, contained n-acetyl-beta-glucosaminidase, n-acetyl-beta-galactosaminidase, beta-galactosidase, and sialidase activities. exoglycosidase activities were found in cell lysates as well as cell-free supernatants from uninfected and wild-type baculo ... | 1993 | 7763590 |
| biotechnological aspects of the production of human pro-kallikrein using the acnpv-baculovirus-expression system. | we have investigated large scale production processes (up to 2 liters) of recombinant proteins using the baculovirus expression system in order to optimize the product yields. experiments using cell lines of spodoptera frugiperda (sf9) and mamestra brassicae (izd-mb0503) were performed to show the different production capacities of the cell lines. the influence of the infection at different cell densities is described. beyond that, tc100-, ipl41- and serum-free ipl41-medium were compared to demo ... | 1993 | 7763638 |
| host-specific transcription of baculovirus genes. | we examined the differential replication of autographa californica multicapsid nuclear polyhedrosis virus (acmnpv; family: baculoviridae) in spodoptera frugiperda (sf) and bombyx mori (bm) cell lines. our previous studies have shown that infection of bm cells is abortive; infectious progeny is not produced, and although virogenic stroma is produced, complete virions or inclusion bodies are not assembled. we designed the present study to determine the extent of infected-cell-specific polypeptides ... | 1993 | 7763702 |
| optimization of growth methods and recombinant protein production in bti-tn-5b1-4 insect cells using the baculovirus expression system. | a novel insect cell line from trichoplusia ni, bti-tn 5b1-4 (tn 5), was compared to spodoptera frugiperda, sf 9, cells for production of two recombinant secreted proteins: truncated epstein-barr viral attachment protein (ebv gp105) and truncated, soluble tissue factor (stf). under optimum conditions for both cell lines, tn 5 cells produced 28-fold more secreted stf than sf 9 cells, respectively, on a per cell basis. the total production of gp105 was similar for the two cell lines. however, tn5 c ... | 1993 | 7764044 |
| fed-batch culture of insect cells: a method to increase the yield of recombinant human nerve growth factor (rhngf) in the baculovirus expression system. | a fed-batch method was developed which increased the density of insect cells (spodoptera frugiperda, sf-9 cells) in suspension culture and the feeding of nutrients improved the yield of a recombinant protein produced by a baculovirus expression system. analysis of spent medium samples indicated that depletions of glucose and glutamine correlated with the retardation of cell growth. feeding of a mixture of nutrients consisting of glucose, glutamine, yeastolate and lipids solution restored the gro ... | 1993 | 7764302 |
| characterization by electrospray mass spectrometry of human ca(2+)-sensitive cytosolic phospholipase a2 produced in baculovirus-infected insect cells. | the 85-kd cytosolic phospholipase a2 (cpla2) is a novel receptor-regulated phospholipase that is thought to initiate the production of inflammatory lipid mediators. since cpla2 is present only in minute amounts (less than 0.01% of total cellular protein) in various cells and tissues, we have used the baculovirus expression system to produce sufficient quantities of cpla2 for structural and functional analysis. the cdna for cpla2 was cloned into a baculovirus expression vector and, upon infection ... | 1994 | 7764328 |
| correct in vivo processing of a chimeric ubiquitin-proapolipoprotein a-i fusion protein in baculovirus-infected insect cells. | the cdna coding for human proapolipoprotein a-i was expressed as a ubiquitin fusion under the control of the polyhedrin promoter in baculovirus-infected sf9 spodoptera frugiperda insect cells. the fusion protein was expressed at high level and was quantitatively cleaved in vivo. the cleaved product was purified and its n-terminal amino acid sequence was established. the data showed that authentic proapolipoprotein a-i has been produced, and thus demonstrated the existence in spodoptera frugiperd ... | 1994 | 7764450 |
| heterologous expression of the plant proteins strictosidine synthase and berberine bridge enzyme in insect cell culture. | the heterologous expression of cdnas encoding the alkaloid biosynthetic enzymes, strictosidine synthase [ec 4.3.3.2] from rauvolfia serpentina and the berberine bridge enzyme [(s)-reticuline: oxygen oxidoreductase (methylene-bridge-forming), ec 1.5.3.9] from eschscholtzia californica, has been achieved in a cell culture (sf9) of the fall army worm, spodoptera frugiperda, using a baculovirus-based expression system. the expression resulted in the overproduction of each plant enzyme in a catalytic ... | 1994 | 7764480 |
| optimization of the growth conditions of sf21 insect cells for high-density perfusion culture in stirred-tank bioreactors. | spodoptera frugiperda insect cells (iplb-sf21-ae) (sf21), infected with baculovirus expression vectors during their exponential growth phase, are commonly used to produce a variety of heterologous recombinant proteins. in the present study the culture conditions of these insect cells were studied to establish high-density suspension cultures with prolonged exponential growth phases. the sf21 cells were grown in 125-ml spinner flasks using five different culture media supplemented with 5% fetal c ... | 1994 | 7764890 |
| clonal variation in the spodoptera frugiperda iplb-sf21-ae insect cell population. | clones have been isolated from the heterogeneous spodoptera frugiperda iplb-sf21-ae insect cell population. five of these clones, in addition to the parent cell line and the sf9 cell line (another clonal isolate of the parent cell line), have been compared in regards to morphology, growth, budded virus synthesis, and recombinant protein synthesis. no significant differences in cell morphology were found among these cell lines. there was, however, a significant difference in the average cell size ... | 1994 | 7764939 |
| continuous insect cell (sf-9) culture with aeration through sparging. | the continuous growth of spodoptera frugiperda sf-9 cells in a 250-ml blown-glass jacketed spinner flask under a direct air sparging environment was investigated. even at 220 ml working volume (about 90% of total volume), this spinner flask provided good mixing and oxygenation as demonstrated by a higher cell density compared with fermentor cultures. this eliminates a common limitation of the traditional spinner flask, namely much lower cell density at high working volume. furthermore, this spin ... | 1994 | 7764966 |
| a prototype recombinant vaccine against respiratory syncytial virus and parainfluenza virus type 3. | we have produced a genetically-engineered chimeric protein composed of the external domains of the respiratory syncytial virus (rsv) fusion (f) protein and the parainfluenza virus type 3 (piv-3) hemagglutinin-neuraminidase (hn) protein in insect cells using the baculovirus expression system. the yield of the soluble chimeric frsv-hnpiv-3 protein could be increased approximately 2-fold by using trichoplasia ni (high five) insect cells in place of spodoptera frugiperda (sf9) for expression. the ch ... | 1994 | 7765021 |
| physiological and environmental factors affecting the growth of insect cells and infection with baculovirus. | insect cell growth can be significantly improved by close attention to the conditions used in the inoculum stages. initial cell concentration, spent medium carry over and inoculum phase withdrawal significantly influenced the growth kinetics of spodoptera frugiperda (sf9) cells. the percentage of cells infected with wild and recombinant baculovirus acnpv and (in the later case) the beta-galactosidase yield in fresh medium was appreciably affected by the stage of the growth curve that cells were ... | 1995 | 7765874 |
| evaluation of monitoring approaches and effects of culture conditions on recombinant protein production in baculovirus-infected insect cells. | the baculovirus infection process of spodoptera frugiperda (sf9) insect cells in oxygen-controlled bioreactors in serum-free medium was investigated using a recombinant autographa californica (acnpv) virus expressing beta-galactosidase enzyme as a model system. a variety of monitoring techniques including trypan blue exclusion, fluorescent dye staining, oxygen uptake rate (our) measurements, and glucose consumption were applied to infected cells to determine the best way of evaluating cell integ ... | 1994 | 7765930 |
| cyp3a4 expressed by insect cells infected with a recombinant baculovirus containing both cyp3a4 and human nadph-cytochrome p450 reductase is catalytically similar to human liver microsomal cyp3a4. | human cytochrome cyp3a4 is the most abundant of all the p450s in human liver and is involved in the metabolism of many environmental toxicants and drugs. kinetic studies with cyp3a4 have been hampered due to low activity of this enzyme obtained from recombinant gene expression systems or difficulty in reconstituting activity with the native enzyme purified from human liver. to overcome these obstacles, we have expressed high levels of catalytically active cyp3a4 and human nadph-cytochrome p450 r ... | 1995 | 7771780 |
| n-glycosylated polypeptides synthesized in the early phase of acnpv infected insect cells. | infected cell specific polypeptides (icsp) in autographa californica nuclear polyhedrosis virus (acnpv) infected spodoptera frugiperda cell line, sf9, were radiolabelled with [14c] mannose and the labelled proteins were monitored at various time intervals. a major glycoprotein of molecular size of 64 kda appears as early as 3h post infection (p.i.) which peaks at 12h p.i., but in presence of tunicamycin this protein is not labelled. two other glycoproteins of molecular size 67 kda and 63 kda app ... | 1995 | 7773189 |
| identification and analysis of a putative origin of dna replication in the choristoneura fumiferana multinucleocapsid nuclear polyhedrosis virus genome. | a recombinant plasmid containing the choristoneura fumiferana multinucleocapsid nuclear polyhedrosis virus (cfmnpv) hindiii r fragment (m.u. 2.2-3.9) was shown to undergo cfmnpv infection-dependent dna replication in cf-124t cells. replication of this dna sequence was detectable by 24 hr p.i. and did not appear to have resulted as a consequence of recombination with the virus genome. replication was inhibited by mimosine, an inhibitor of eukaryotic dna replication. these data suggest that hindii ... | 1995 | 7778276 |
| the wild-type autographa californica nuclear polyhedrosis virus induces apoptosis of spodoptera littoralis cells. | spodoptera littoralis cells infected with the autographa californica multiple nuclear polyhedrosis virus (acmnpv) yielded significantly lower budded virus titers than spodoptera frugiperda-infected cells and produced very low levels of polyhedrin. relative to acmnpv-infected s. frugiperda sf9 cells viral dna replication was severely reduced in spodoptera littoralis sl2 cells. microscopic examination of sl2-infected cells revealed progressive cell blebbing starting at 6-8 hr postinfection and cul ... | 1995 | 7778284 |
| nucleotide sequence and transcriptional analysis of the gp41 gene of spodoptera frugiperda nuclear polyhedrosis virus. | the spodoptera frugiperda multiple nucleocapsid nuclear polyhedrosis virus (sfmnpv) gp41 structural protein gene was located in the 1.9 kbp ecori-s fragment and sequenced. an open reading frame (orf) of 999 nucleotides was detected that encoded a protein of 332 amino acids. the gp41 gene transcript was detected after 12 h post-infection (p.i.) and remained detectable at 48 h p.i. two major mrnas, about 1.6 and 2.8 kb in length, were determined by northern blot analysis. primer extension analysis ... | 1995 | 7782772 |
| mutations in the autographa californica multinucleocapsid nuclear polyhedrosis virus 25 kda protein gene result in reduced virion occlusion, altered intranuclear envelopment and enhanced virus production. | serial passage of nuclear polyhedrosis viruses (npvs) through cultured cell lines results in the appearance of mutants with a complex phenotype referred to as the 'few polyhedra' (fp) phenotype. the altered plaque morphology and reduced occlusion production associated with the fp phenotype have been observed in autographa californica multinucleocapsid nuclear polyhedrosis virus (acmnpv) bearing mutations in the gene encoding the 25 kda protein (25k gene). in this study, we sequenced the 25k gene ... | 1995 | 7782773 |
| potassium channel structure and function as reported by a single glycosylation sequon. | inwardly rectifying k+ channels (irks) are highly k(+)-selective, integral membrane proteins that help maintain resting the membrane potential and cell volume. integral membrane proteins as a class are frequently n-glycosylated with the attached carbohydrate being extracellular and perhaps modulating function. however, dynamic effects of glycosylation have yet to be demonstrated at the molecular level. romk1, a member of the irk family is particularly suited to the study of glycosylation because ... | 1995 | 7797521 |
| preparation and characterization of human rheumatoid arthritic synovial fluid phospholipase a2 produced by recombinant baculovirus-infected insect cells. | we prepared human rheumatoid arthritic synovial fluid phospholipase a2 (pla2) [ec 3.1.1.4] from insect cells infected with a recombinant baculovirus. the pla2 dna was designed, changing the original codons to those used frequently in the polyhedrin gene. sixteen oligo-deoxynucleotides ranging from 40 to 70 nucleotides were chemically synthesized and then assembled to form the whole pla2 gene. the gene thus synthesized was then placed under the control of the polyhedrin promoter of autographa cal ... | 1994 | 7798190 |
| kinetic properties of the alpha 2 beta 1 and alpha 2 beta 2 isozymes of the na,k-atpase. | the presence of multiple isoforms of the alpha and beta subunits of the na,k-atpase in most mammalian tissues has hindered the understanding of the roles of the individual isoforms in directing na,k-atpase function. expression of the na,k-atpase subunits in insect cells using recombinant baculoviruses has proven to be a useful system for the study of na,k-atpase function. using this system, we have expressed the rat na,k-atpase alpha 2 beta 1 and alpha 2 beta 2 isoforms in sf-9 insect cells, a c ... | 1995 | 7819213 |
| characterization of an insect cell-derived theileria parva sporozoite vaccine antigen and immunogenicity in cattle. | previous data showed that six out of a group of nine cattle inoculated with ns1-p67, a recombinant form of a 67-kda theileria parva sporozoite surface protein, were immune to east coast fever. this bacterially expressed antigen encoded all 709 amino acid residues of p67 fused to the c-terminal end of 87 residues derived from ns1, a structural protein of influenza virus, and a linker dna sequence. ns1-p67 lacked reactivity with tpm 12, a monoclonal antibody to native p67, and had an estimated mol ... | 1995 | 7822014 |
| protective efficacy in mice of a secreted form of recombinant dengue-2 virus envelope protein produced in baculovirus infected insect cells. | we constructed a recombinant baculovirus encoding a dengue (den)-2 virus envelope glycoprotein truncated of 102 amino acids (aa) at its c-terminus (d2e delta 102). the production, processing and transportation of the recombinant protein in baculovirus-infected spodoptera frugiperda (sf9) cells and its immunogenic properties in mice were compared to those of a previously characterized recombinant den-2 e-protein with a 71aa c-terminal truncation (d2e delta 71). both proteins were transported thro ... | 1994 | 7826210 |
| altered glycosylation and selected mutation in recombinant human complement component c9: effects on haemolytic activity. | recombinant wild-type and mutated forms of human complement component c9 have been synthesized in baculovirus-infected insect cells. wild-type recombinant c9 was indistinguishable from native c9, as judged by haemolytic activity, trypsin and alpha-thrombin digestion, reaction with antibodies to c9, enzymatic deglycosylation to the same core size and polymerization in the presence of zn2+. replacement of the native signal peptide with the honey-bee melittin signal peptide, and replacement of spod ... | 1994 | 7835977 |
| plaque assay and replication of tipula iridescent virus in spodoptera frugiperda ovarian cells. | a plaque assay was developed for the study of tipula iridescent virus (tiv) replication using a cell line derived from the fall army worm spodoptera frugiperda (sf9). infection and plaque formation were monitored with time by phase contrast microscopy, video and fluorescent light microscopy. structure of virions, viroplasmic centres and organelles of infected cells were examined by transmission electron microscopy (tem). after 4 h postinfection, plaques were visibly detected within the cell mono ... | 1994 | 7839010 |
| functional chicken gizzard heavy meromyosin expression in and purification from baculovirus-infected insect cells. | the heavy chain and the essential and the regulatory light chains of chicken gizzard heavy meromyosin (hmm) were coexpressed in spodoptera frugiperda (fall armyworm) cells infected with a mixture of two recombinant autographa californica baculoviruses. soluble hmm consisting of the heavy chain and the two types of light chains was obtained. the recombinant hmm was purified from the virus-infected cells and characterized. the regulatory light chain of the isolated recombinant hmm was phosphorylat ... | 1995 | 7846039 |
| signal transduction in sf9 insect cells: endocytosis of recombinant cd4 after phorbol ester treatment. | in spodoptera frugiperda (sf9) insect cells infected with a recombinant baculovirus carrying a gene construct encoding human cd4 endocytosis of cd4 is induced after stimulation with phorbol-12-myristate-13-acetate (pma). stimulation of endocytosis with pma reduced the amount of full-length cd4 on the plasma membrane of sf9 cells by 50% after 2 hours. endocytosis of cd4 is blocked after intracellular delivery by cationic liposomes of a monoclonal antibody directed against a cytoplasmic sequence o ... | 1995 | 7857297 |
| human pancreatic procolipase expressed in insect cells: purification and characterization. | a cdna clone encoding human pancreatic procolipase was incorporated into a recombinant baculovirus. spodoptera frugiperda insect cells infected with the recombinant baculovirus secreted procolipase into the medium, which could be isolated in a single step by immunoaffinity chromatography. the highly purified protein reactivated human pancreatic lipase in a concentration-dependent fashion and was efficiently converted to colipase by limited trypsin digestion. this expression system is suitable fo ... | 1994 | 7858428 |
| effects of co-expressing chaperone bip on functional antibody production in the baculovirus system. | the assembly pathway of the insect cell spodoptera frugiperda (sf-9) was engineered to include expression of the murine chaperone immunoglobulin heavy chain binding protein (bip) using the baculovirus vector. the impact of bip coexpression on the production and secretion of functional and soluble recombinant immunoglobulin igg levels was evaluated. recombinant bip was found to associate specifically with immunoglobulins in immunoprecipitation studies. coinfection of insect cells with a bip-conta ... | 1994 | 7858430 |
| the structure and function of the first component of complement: genetic engineering approach (a review). | the availability of cdna and genomic clones for the subcomponents of c1, as well as the recognition of the modular organization of serine-proteases have opened up exciting new possibilities for approaching structural problems. in this review the latest achievements of combined protein engineering, functional and structural studies are summarized. the concept of this research is to construct deletion, point and hybrid mutants of the highly homologous c1r and c1s subcomponents, to reveal the funct ... | 1994 | 7866721 |
| production of measles nucleoprotein in different expression systems and its use as a diagnostic reagent. | measles nucleoprotein has been successfully expressed in three different hosts, bacterial (escherichia coli bl21 de3), insect (spodoptera frugiperda; sf9) and mammalian (primary human fibroblasts) cells, each producing an antigenic protein of m(r) 60 kda. the nucleoprotein produced in all three hosts was used in an elisa for the detection of antibodies to measles virus in a cohort of haemagglutinin-positive or -negative human sera. data produced from baculovirus and adenovirus-based antigens ind ... | 1994 | 7868644 |
| agonist-dependent phosphorylation of human muscarinic receptors in spodoptera frugiperda insect cell membranes by g protein-coupled receptor kinases. | agonist-dependent phosphorylation of g protein-coupled receptors (gprs) by g protein-coupled receptor kinases (grks) is proposed to be a key event initiating homologous receptor desensitization. a technical limitation hindering identification of gprs as grk substrates has been the necessity to use purified and reconstituted receptors in grk assays. here, the human m2 and human m3 (hm3) muscarinic cholinergic receptors (machrs), which couple to attenuation of adenylyl cyclase and stimulation of p ... | 1995 | 7870029 |
| proteolysis of p6.9 induced by cytochalasin d in autographa californica m nuclear polyhedrosis virus-infected cells. | cytochalasin d, a fungus-derived compound that interferes with actin polymerization, inhibits autographa californica m nuclear polyhedrosis virus production in infected spodoptera frugiperda (iplb-sf-21) cells. cytochalasin d appears to inhibit nucleocapsid morphogenesis by interfering with nucleoprotein packaging. we were interested in determining, therefore, whether the drug affected the synthesis or processing of p6.9, the major core protein involved in nucleoprotein packaging. we found that ... | 1995 | 7871717 |
| purification and properties of a golgi-derived (alpha 1,2)-mannosidase-i from baculovirus-infected lepidopteran insect cells (iplb-sf21ae) with preferential activity toward mannose6-n-acetylglucosamine2. | because the availability and subcellular distribution of processing mannosidases in cells play such powerful roles in determining ultimate structures of glycoconjugates, we desired to identify, characterize, and investigate possible regulation of mannosidases in infected and noninfected lepidopteran insect cells. since our previous observations that a mannosidase activity that converted man6glcnac2 to man5glcnac2 was enhanced in virus-infected cells, thus providing the necessary intermediate for ... | 1995 | 7873528 |
| overexpression and characterization of the human peroxisomal acyl-coa oxidase in insect cells. | human liver peroxisomes contain two acyl-coa oxidases, namely, palmitoyl-coa oxidase and a branched chain acyl-coa oxidase. the palmitoyl-coa oxidase (acox) oxidizes the coa esters of straight chain fatty acids and prostaglandins and donates electrons directly to molecular oxygen, thereby producing h2o2. the inducibility of this h2o2-generating acox in rat and mouse liver by peroxisome proliferators and the postulated role of the resulting oxidative stress in hepatocarcinogenesis generated inter ... | 1995 | 7876265 |
| identification and characterization of a second putative origin of dna replication in a baculovirus of orgyia pseudotsugata. | a 7.5-kb region (96.8-2.5 m.u.), called op5, of the orgyia pseudotsugata multinucleocapsid nuclear polyhedrosis virus (opmnpv) genome that contains an origin of dna replication was characterized. this region replicates several times more efficiently and is unrelated to the previously identified putative origin of replication located on the viral hindiii-n fragment. in contrast to hindiii-n, the origin on op5 contains repeated sequences with limited sequence identity to the homologous regions fro ... | 1995 | 7886962 |
| expression and characterization of the alpha 2 subunit of the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (ampa)-selective glutamate receptor channel in a baculovirus system. | using a baculovirus expression vector system, the alpha 2 subunit of the mouse alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (ampa)-selective glutamate receptor (glur) channel was expressed in spodoptera frugiperda insect cells. immunoblotting using the antibody made to the synthetic peptide corresponding to the c-terminus of glur alpha 2 and [35s]methionine/[35s]cysteine metabolic radiolabeling revealed the major 102-kda and the minor 98-kda protein bands. metabolic radiolabeling with t ... | 1994 | 7889366 |
| characterization of wild-type and amyotrophic lateral sclerosis-related mutant cu,zn-superoxide dismutases overproduced in baculovirus-infected insect cells. | we describe the use of a baculovirus expression system to overproduce human cu,zn-superoxide dismutase (sod). spodoptera frugiperda (sf21) insect cells infected with a baculovirus carrying the cu,zn-sod cdna synthesized a large amount of cu,zn-sod apoprotein in the conventional medium. the sod activity of the apoprotein, which was initially very low, increased in a dose-dependent manner when cu2+ and zn2+ were added to the medium. cells grown in media supplemented with cu2+ alone exhibited nearl ... | 1995 | 7891072 |
| expression and characterization of functional bovine cation-dependent mannose 6-phosphate receptors in baculovirus-infected insect cells. | a recombinant baculovirus containing the cdna for the full-length bovine cation-dependent mannose 6-phosphate receptor (cd-mpr) was generated by homologous recombination. spodoptera frugiperda (sf9) and trichoplusia ni 5b1-4 (high five) insect cells, which do not contain endogenous mprs as determined by western blot analyses and pentamannosyl phosphate-agarose affinity chromatography, were infected with recombinant baculovirus. the infected cells expressed 26-, 29-, 32-, 35-, and 39-kda species ... | 1995 | 7893168 |
| primary structure of a folate-dependent trifunctional enzyme from spodoptera frugiperda. | the dehydrogenase and synthetase activities of the nadp-dependent methylenetetra-hydrofolate dehydrogenase-cyclohydrolase-synthetase are undetectable in extracts of the spodoptera frugiperda cell line, sf9. however, a single cdna encoding this protein was isolated from a library and sequenced. the deduced amino acid sequence codes for a protein of 933 amino acids in length that shows 59% identity to the human enzyme. the cdna inserted in the yeast expression vector pvt102-u complements a purine ... | 1995 | 7893749 |
| expression of the human interleukin-2 receptor gamma chain in insect cells using a baculovirus expression vector. | the gene encoding the gamma-chain of the human interleukin-2 receptor was expressed in lepidopteran insect cells using the baculovirus expression vector system. the corresponding gene was inserted under the polyhedrin promoter of the autographa californica nuclear polyhedrosis virus and expressed in the spodoptera frugiperda insect cell line sf9 during viral infection. the recombinant receptor protein was identified by immunoblotting in cell lysates, prepared from insect cells infected with the ... | 1995 | 7899821 |
| baculovirus-mediated expression of the epidermal growth factor-like modules of human factor ix fused to the factor xiiia transamidation site in fibronectin. evidence for a direct interaction between the nh2-terminal epidermal growth factor-like module of factor ixa beta and factor x. | factor ix is a vitamin k-dependent procoagulant zymogen of a serine protease. in the presence of ca2+ the active form of factor ix (factor ixa beta) forms a complex with factor viiia on suitable phospholipid surfaces such as aggregated platelets. this macromolecular complex rapidly activates factor x. we have previously provided data that suggest an interaction between the nh2-terminal epidermal growth factor (egf)-like module of factor ixa beta and the substrate factor x. in an alternative appr ... | 1994 | 7906269 |
| enzymatic catalysis of disulfide formation. | the formation of disulfide bridges in membrane and secretory proteins occurs during the protein folding process in the endoplasmic reticulum of eukaryotic cells and the periplasm of prokaryotes. the formation of disulfide bridges is facilitated by the thiol-disulfide oxidoreductases, protein disulfide isomerase (pdi) in eukaryotes and dsba in prokaryotes. structure-function analysis of these soluble proteins demonstrates that their active sites are sequences with similarity to the active site of ... | 1994 | 7909462 |
| similar ligand binding in recombinant human alpha 2 c2-adrenoceptors produced in mammalian, insect and yeast cells. | ligand binding properties were investigated in recombinant human alpha 2c2-adrenoceptors expressed in three different host systems: shionogi s115 mouse mammary tumour cells, spodoptera frugiperda sf9 insect cells and saccharomyces cerevisiae yeast cells. the expected 43 kda alpha 2c2 protein was visualized with immunoblotting using a polyclonal alpha 2c2-receptor antibody. [3h]rauwolscine binding in cell homogenates or membranes (bmax 3-11 pmol/mg protein; kd approximately 5.5 nm) was inhibited ... | 1994 | 7911432 |
| human 17 beta-hydroxysteroid dehydrogenase: overproduction using a baculovirus expression system and characterization. | estrogenic 17 beta-hydroxysteroid dehydrogenase (17 beta-hsd) plays a pivotal role in the synthesis of estrogens. we overproduced human placental estrogenic 17 beta-hsd using a baculovirus expression system for the study of the enzyme mechanism. a cdna encoding the entire open reading frame of human 17 beta-hsd was inserted into the genome of autographa californica nuclear polyhedrosis virus and expressed in spodoptera frugiperda (sf9) insect cells. metabolic labeling and western blot analysis u ... | 1994 | 7918113 |
| synapsin iia bundles actin filaments. | synapsins are neuron-specific phosphoproteins associated with small synaptic vesicles in the presynaptic nerve terminal. synapsin i, which has been demonstrated to bundle f-actin in vitro, has been postulated to regulate neurotransmitter release by cross-linking synaptic vesicles to the actin cytoskeleton. to investigate the possible interaction of synapsin ii with actin filaments, we expressed synapsin ii in spodoptera frugiperda and high five insect cells using a recombinant baculovirus. purif ... | 1994 | 7931311 |
| expression of functional metabotropic and ionotropic glutamate receptors in baculovirus-infected insect cells. | an ionotropic glutamate receptor of the kainate subtype (glur6) and a g-protein coupled metabotropic glutamate receptor (mglur1 alpha) were expressed and studied in two insect cell lines: sf9 cells from spodoptera frugiperda and mg1 cells from trichoplusia ni. application of kainate to glur6-infected mg1 cells produced kainate-activated currents. glutamate activation of mglur1 alpha in mg1- and sf9-infected cells caused rapid, transient increases in intracellular calcium levels. this effect was ... | 1994 | 7936400 |
| cystine-rich fish antifreeze is produced as an active proprotein precursor in fall armyworm cells. | recombinant cystine-rich fish antifreeze protein (afp) was produced by fall armyworm cells from a baculovirus expression vector containing sea raven afp cdna. the natural signal sequence encoded in the cdna directed secretion of the antifreeze into the medium, from where it was recovered and purified to homogeneity. the m(r) of the exported protein (16k), as determined by sds-page, was larger than that (14k) of mature afp isolated from sea raven serum. sequencing of the recombinant afp showed th ... | 1994 | 7945337 |
| expression of a cannabinoid receptor in baculovirus-infected insect cells. | a cannabinoid receptor recombinant baculovirus (acnpv-thcr) has been constructed and employed to express rat neural cannabinoid receptors. northern analysis of total rna from spodoptera frugiperda (sf9) insect cells infected with acnpv-thcr revealed novel hyper-production of a 3.3 kb transcript when probed with nick-translated rat cannabinoid receptor cdna. optimal viral protein expression was observed in 35s-metabolically labeled acnpv-thcr-infected sf9 cells at a multiplicity of infection of 2 ... | 1994 | 7945417 |
| cloning and expression of a soluble sialidase from chinese hamster ovary cells: sequence alignment similarities to bacterial sialidases. | a cdna encoding a soluble sialidase from chinese hamster ovary (cho) cells has been cloned and expressed. completely degenerate oligonucleotide primers, which were based on the amino acid sequence of peptides obtained from the purified sialidase (warner et al., glycobiology, 3, 455-463, 1993), and the polymerase chain reaction, with single-stranded cdna template, were employed to generate a unique oligonucleotide probe. the unique probe of 93 bp was used for screening a lambda gt 10 cho cell cdn ... | 1994 | 7949662 |
| microtubule-associated protein function: lessons from expression in spodoptera frugiperda cells. | the phenotypes induced by the expression of neuronal microtubule-associated proteins (maps) in sf9 cells have provided data on the in situ function of these proteins. both map2 and tau can induce long processes in sf9 cells, and the processes contain bundles of microtubules. in both cases the microtubules are aligned with their plus ends distal. tau expression usually induces a single process that is unbranched and of uniform caliber. processes can form even when the cells are grown in suspensio ... | 1994 | 7954847 |
| apoptosis of spodoptera frugiperda cells induced by okadaic acid is abrogated by baculovirus infection. | okadaic at low concentrations specifically inhibits protein phosphatase 2a. treatment of cells with this inhibitor leads to growth arrest and cell injury within 1 to 2 days. after testing several different types of cells we found that mouse 3t6 cells and insect sf9 cells responded to okadaic acid by apoptosis, as defined by changes in morphology and degradation of chromatin to fragments of nucleosome size. infection of insect cells by baculovirus prevented the okadaic acid-induced apoptosis. ana ... | 1994 | 7957672 |
| overproduction and purification of mycobacterium tuberculosis chaperonin 10. | the overexpression of the mycobacterium tuberculosis chaperonin 10 (cpn10)-encoding gene was accomplished using baculovirus expression vectors. the product was immunoreactive with a cpn10 monoclonal antibody (mab) and had an electrophoretic mobility identical to authentic cpn10. the baculovirus system was most successful in terms of reaching nearly the full expression potential of the system. recombinant cpn10 was purified from recombinant baculovirus-infected spodoptera frugiperda cells by isoe ... | 1994 | 7959041 |