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molecular cloning and nucleotide sequence of the protyrosinase gene, melo, from aspergillus oryzae and expression of the gene in yeast cells.the coding region of the protyrosinase gene, melo, from aspergillus oryzae occupies 1671 base pairs of the genomic dna and is separated into two exons by one intron. the full-length cdna of the melo gene was cloned. analysis of the 1617 base pairs nucleotide sequence revealed a single open reading frame coding 539 amino acid residues. the cdna has been expressed in yeast cells. the predicted protein product derived from the melo gene is identified by western blotting and activity determination. ...19957893753
ethylene oxide: induction of specific-locus mutations in the ad-3 region of heterokaryon 12 of neurospora crassa and implications for genetic risk assessment of human exposure in the workplace.ethylene oxide (eto) is an important industrial intermediate used extensively in the production of ethylene glycol, as a fumigant, and as a sterilant of choice for various medical devices. the mutagenicity of eto was studied for the induction of specific-locus mutations in the adenine-3 (ad-3) region of a two-component heterokaryon (h-12) of neurospora crassa. the objectives of these studies with eto were to rank its mutagenic potency and to compare its mutational spectrum for induced specific-l ...19957898502
molybdenum cofactor biosynthesis in neurospora crassa: biochemical characterization of pleiotropic molybdoenzyme mutants nit-7, nit-8, nit-9a, b and c.available mutants of molybdenum cofactor (moco) biosynthesis of neurospora crassa were studied for converting factor activity and for in vitro molybdate repair of nitrate reductase (nr) activity. mutant nit-7 was found to contain an activity that fits the functional definition of converting factor activity in escherichia coli. its high molecular weight fraction converts a low molecular weight compound from nit-1 and nit-8 into biologically active molybdopterin (mpt). like nit-1, mutant nit-8 is ...19957899494
phylogenetic analysis of ten black yeast species using nuclear small subunit rrna gene sequences.the nuclear small subunit rrna genes of authentic strains of the black yeasts exophiala dermatitidis, wangiella dermatitidis, sarcinomyces phaemuriformis, capronia mansonii, nadsoniella nigra var. hesuelica, phaeoannellomyces elegans, phaeococcomyces exophialae, exophiala jeanselmei var. jeanselmei and e. castellanii were amplified by pcr and directly sequenced. a putative secondary structure of the nuclear small subunit rrna of exophiala dermatitidis was predicted from the sequence data. alignm ...19958526477
isolation and characterization of the catalytic subunit of protein phosphatase 2a from neurospora crassa.the catalytic subunit of protein phosphatase 2a (pp2ac) was purified from neurospora crassa extract by (nh4)2so4-ethanol precipitation followed by deae-sephacel, heparin-sepharose, and monoq chromatography steps about 900-fold to a specific activity of 1200 u/g with a 2% yield. the apparent m(r) of pp2ac was estimated to be 35 kda by gel filtration and 33 kda by sds polyacrylamide gel electrophoresis. half maximal inhibition of pp2ac was achieved at 0.3 nm okadaic acid, 0.1 nm microcystin-lr, 56 ...19958529028
detection of additional restriction fragment length polymorphisms among the weakly virulent (nonaggressive) and highly virulent (aggressive) isolates of leptosphaeria maculans.isolates of leptosphaeria maculans were analyzed for their genetic relatedness based on dna restriction fragment length polymorphisms (rflps), employing as southern hybridization probes a combination of heat shock responsive genes (hsp70 and hsp80 from neurospora crassa), the cutinase gene of magnaporthe grisea, and cloned genomic dna sequences from a virulent strain. southern hybridization analysis revealed a high frequency of dna polymorphism. restriction fragments generated by each enzyme-pro ...19958542555
the roles of ca2+ and plasma membrane ion channels in hyphal tip growth of neurospora crassa.growing hyphae of the ascomycete fungus neurospora crassa contained a tip-high gradient of cytoplasmic ca2+, which was absent in non-growing hyphae and was insensitive to gd3+ in the medium. patch clamp recordings in the cell-attached mode, from the plasma membrane of these hyphae, showed two types of channel activities; spontaneous and stretch activated. the spontaneous channels were identified as inward k+ channels based on inhibition by tetraethylammonium. the stretch activated channels had i ...19958586653
the novel nuclear gene dss-1 of saccharomyces cerevisiae is necessary for mitochondrial biogenesis.a previously unknown nuclear gene dss-1 from saccharomyces cerevisiae was cloned and sequenced. the gene was isolated as a multicopy suppressor of a disruption of the suv-3 gene coding for a dead/h box protein involved in processing and turnover of mitochondrial transcripts. the dss-1 gene codes for a 970 amino-acid protein of molecular weight 111 kda and is necessary for mitochondrial biogenesis. amino-acid sequence analysis indicates the presence of motifs characteristic for escherichia coli r ...19958590460
expression of the neurospora crassa metallothionein gene in escherichia coli and its effect on heavy-metal uptake.the gene coding for the neurospora crassa metallothionein protein was chemically synthesized and cloned into the fusion expression vectors pmal-c and pmal-p. cell-fractionation experiments demonstrated the proper localization of the pmal-c and pmal-p- expressed proteins to the cytosol and periplasm of the bacteria respectively. control bacteria as well as the recombinant bacteria producing the metallothionein protein were incubated with solutions of 109cd at concentrations of 0.2 microm, 1 micro ...19958590662
mitochondrial receptor complex from neurospora crassa and saccharomyces cerevisiae. 19958592451
generation and characterization of nadh: ubiquinone oxidoreductase mutants in neurospora crassa. 19958592454
different respiratory-defective phenotypes of neurospora crassa and saccharomyces cerevisiae after inactivation of the gene encoding the mitochondrial acyl carrier protein.the nuclear genes (acp-1, acp1) encoding the mitochondrial acyl carrier protein were disrupted in neurospora crassa and saccharomyces cerevisiae. in n. crassa acp-1 is a peripheral subunit of the respiratory nadh : ubiquinone oxidoreductase (complex i). s. cerevisiae lacks complex i and its acp1 appears to be located in the mitochondrial matrix. the loss of acp-1 in n. crassa causes two biochemical lesions. firstly, the peripheral part of complex i is not assembled, and the membrane part is not ...19958595652
the orientation of gene maps by recombination of flanking markers for the am locus of neurospora crassa.fincham (1967), smyth (1973 b) and rambosek and kinsey (1983) have each generated fine-structure maps of the am gene of neurospora crassa. each map had a consistent linear order of alleles but the assignment of an orientation with respect to other linkage group-v loci differed. fincham found the end marked by the am6 allele to be at the distal end of the locus, smyth found am6 to be at the proximal end while the data of rambosek and kinsey did not suggest an orientation. smyth s orientation has ...19958595655
functional molecular aspects of the nadh dehydrogenases of plant mitochondria.there are multiple routes of nad(p)h oxidation associated with the inner membrane of plant mitochondria. these are the phosphorylating nadh dehydrogenase, otherwise known as complex i, and at least four other nonphosphorylating nad(p)h dehydrogenases. complex i has been isolated from beetroot, broad bean, and potato mitochondria. it has at least 32 polypeptides associated with it, contains fmn as its prosthetic group, and the purified enzyme is sensitive to inhibition by rotenone. in terms of su ...19958595975
mutational analysis reveals dispensability of the n-terminal region of the aspergillus transcription factor mediating nitrogen metabolite repression.mutational analysis has enabled identification and localization of an upstream exon of the area gene of aspergillus nidulans mediating nitrogen metabolite repression. a mutation in the initiation codon and frameshift mutations, which revert by restoration of the reading frame, established the coding role of the exon and mutations affecting intron splicing in conjunction with dna sequencing of reverse transcriptase polymerase chain reaction (rt-pcr) products localized the coding region intron. th ...19958596437
identification and expression of the neurospora crassa mei-3 gene which encodes a protein homologous to rad51 of saccharomyces cerevisiae.the mei-3 gene of neurospora crassa encodes a homolog of the escherichia coli reca and saccharomyces cerevisiae rad51 proteins, which are required for recombination and repair of dna double-strand breaks. to determine the molecular function of mei3 protein, anti-mei3 antibody was prepared and used in western blot analysis. the antibody cross-reacted only with crude extracts prepared from perithecia, the fruiting bodies of neurospora. the molecular weight of the mei3 protein was estimated to be 3 ...19958552049
functional analysis by site-directed mutagenesis of individual amino acid residues in the flavin domain of neurospora crassa nitrate reductase.nitrate reductase of neurospora crassa is a complex multi-redox protein composed of two identical subunits, each of which contains three distinct domains, an amino-terminal domain that contains a molybdopterin cofactor, a central heme-containing domain, and a carboxy-terminal domain which binds a flavin and a pyridine nucleotide cofactor. the flavin domain of nitrate reductase appears to have structural and functional similarity to ferredoxin nadph reductase (fnr). using the crystal structure of ...19958552051
control of neurospora crassa morphology by cyclic adenosine 3', 5'-monophosphate and dibutyryl cyclic adenosine 3', 5'-monophosphate.the role of cyclic adenosine 3', 5'-monophosphate (cyclic amp) in the control of the neurospora asexual life cycle was studied. endogenous cyclic amp levels were 10 to 20 times higher in strains having the wild-type cr-1 allele than in those carrying the mutated allele. in a wild-type strain these levels remained constant throughtout the entire growth period in shaken liquid cultures, except during a short period at the beginning of the stationary growth phase. in this period a marked increase i ...1995177398
isolation and characterization of nuclei from neurospora crassa.a procedure was developed for isolating nuclei from either the conidial or germinated conidial growth phase of neurospora crassa. a frozen conidial suspension was lysed by passage through a french pressure cell, and the nuclei were freed from the broken cells by repeated homogenization in an omni-mixer. pure nuclei were obtained from the crude nuclear fraction by density banding in a ludox gradient. the final nuclear yield was 20 to 30%. the nuclei had a deoxyribonucleic acid (dna):ribonucleic a ...1995162536
nuclear migration advances in fungi.nuclear migration encompasses three areas: separation of daughter nuclei during mitosis, congress of parental nuclei before they fuse during fertilization, and positioning of nuclei in interphase cells. this review deals primarily with interphase nuclear migration, which is crucial for events as disparate as vertebrate embryonic development and growth of fungal mycelia. mutants of aspergillus nidulans, neurospora crassa and saccharomyces cerevisiae have been particularly informative, and a detai ...199514732112
primer sets developed to amplify conserved genes from filamentous ascomycetes are useful in differentiating fusarium species associated with conifers.we examined the usefulness of primer sets designed to amplify introns within conserved genes in filamentous ascomycetes to differentiate 35 isolates representing six different species of fusarium commonly found in association with conifer seedlings. we analyzed restriction fragment length polymorphisms (rflp) in five amplified pcr products from each fusarium isolate. the primers used in this study were constructed on the basis of sequence information from the h3, h4, and (beta)-tubulin genes in ...199516534991
partial amino acid sequences of peptidyl-prolyl isomerases offusarium sporotrichioides.peptidyl-proprylyl cis-trans isomerase (ppiase) activity was observed from crude extract of fusarium sporotrichioides. proteins from this fungi were separated by two-dimensional polyacrylamide gel electrophoresis and more than one thousand protein spots were separated. two cytosolic ppiases were found by the n-terminal sequencing from the two separated spots. the n-terminal 41 residues of the major protein spot showed high sequence identity (63.4%) with ppiase from neurospora crassa. this protei ...199511725072
only the mature form of the plastidic chorismate synthase is enzymatically active.coding regions of a cdna for precursor and mature chorismate synthase (cs), a plastidic enzyme, from corydalis sempervirens were expressed in escherichia coli as translational fusions to glutathione-s-transferase. fusion proteins were purified, and precursor and mature forms of cs were then released by proteolytic cleavage with factor xa. although mature cs was enzymatically active after release, activity could be detected neither for the precursor cs nor for corresponding glutathione-s-transfer ...199512228531
effects of temperature shifts on the metabolism of trehalose in neurospora crassa wild type and a trehalase-deficient (tre) mutant. evidence against the participation of periplasmic trehalase in the catabolism of intracellular trehalose.the effects of temperature shifts on the metabolism of trehalose in neurospora crassa were studied in conidiospore germlings of a wild type strain, and of a mutant (tre), deficient in the activity of periplasmic trehalase. when the temperature of the medium was raised from 30 degrees c to 45 degrees c both strains accumulated trehalose, either in media supplemented with glucose or with glycerol as carbon sources. the profiles of glycolysis metabolites suggested that at 45 degrees c glycolysis wa ...19958541310
mutations affecting the biosynthesis of s-adenosylmethionine cause reduction of dna methylation in neurospora crassa.a temperature-sensitive methionine auxotroph of neurospora crassa was found in a collection of conditional mutants and shown to be deficient in dna methylation when grown under semipermissive conditions. the defective gene was identified as met-3, which encodes cystathionine-gamma-synthase. we explored the possibility that the methylation defect results from deficiency of s-adenosylmethionine (sam), the presumptive methyl group donor. methionine starvation of mutants from each of nine complement ...19958532524
sequence repeat-induced disruption of the major heat-inducible hsp70 gene of neurospora crassa.the process of repeat-induced point mutation (rip) was used to disrupt hsps-1, the gene encoding the major heat-inducible member of the hsp70 family of neurospora crassa. a plasmid dna, containing an incomplete copy of hsps-1 and the selectable marker qa-2+, was introduced into germinated conidia. the sexual progeny of transformants with ectopically integrated hsps-1 dna was examined for rip by southern-blot analysis of mboi- and sau3a-digested genomic dna. progeny strains, showing rip, were tes ...19958595654
a suppressor mutation which suppresses adenylyl cyclase mutations in neurospora crassa.a spontaneous suppressor mutant, hah, which suppressed the colonial growth of adenylyl cyclase mutants (cr-1) was isolated. the morphology of cr-1 hah was filamentous, but slightly different from that of wild type on solid medium. the hah strain formed many high aerial hyphae, but did not form any conidia. the expression levels of an adenylyl cyclase gene, nac, in both hah and cr-1 hah were much higher than those in wild type or in cr-1. the level of camp in cr-1 was very low but returned to clo ...19958574907
osmotic effects on the polyamine pathway of neurospora crassa.in bacteria, mammals, and certain plants, the induction of the polyamine synthetic enzyme, ornithine decarboxylase (odc), and the accumulation of its product, putrescine, follows osmotic manipulations of cells. in at least some of these cases, this response is indispensable for survival. we wished to determine whether the polyamine pathway of neurospora crassa was regulated in response to hyper- or hypoosmotic conditions. unlike odc of most other classes of organisms, the n. crassa enzyme and th ...19958574906
cytogenetics of an intrachromosomal transposition in neurospora.knowledge of intrachromosomal transpositions has until now been primarily cytological and has been limited to drosophila and to humans, in both of which segmental shifts can be recognized by altered banding patterns. there has been little genetic information. in this study, we describe the genetic and cytogenetic properties of a transposition in neurospora crassa. in tp(ir-->il)t54m94, a 20 map unit segment of linkage group i has been excised from its normal position and inserted near the centro ...19958565702
regulation of the expression of three housekeeping genes encoding subunits of the neurospora crassa vacuolar atpase.the vacuolar atpase is a complex enzyme and is encoded by at least nine genes, which appear to be scattered throughout the genome. we have examined the vma-1 vma-2, and vma-3 genes, which encode subunits present in multiple copies within the neurospora crassa vacuolar atpase. we wished to see if the expression of these genes is coordinately regulated and if these genes contain similar promoter elements. a region was sequenced of approximately 1 kb located upstream of the protein coding region fo ...19957500957
purification of a heteromeric ccaat binding protein from neurospora crassa.expression of the neurospora crassa am (nadp-specific glutamate dehydrogenase) gene is controlled by two upstream enhancer-like elements designated ursam alpha and ursam beta. ursam alpha is localized between - 1.3 and - 1.4 kb with respect to the major transcriptional start site. deletion of a 90 bp sequence containing this element resulted in the loss of approximately 50% of normal glutamate dehydrogenase expression. gel mobility shift analysis indicated that a nuclear protein from neurospora ...19957500955
the neurospora organelle motor: a distant relative of conventional kinesin with unconventional properties.the "conventional" kinesins comprise a conserved family of molecular motors for organelle transport that have been identified in various animal species. organelle motors from other phyla have not yet been analyzed at the molecular level. here we report the identification, biochemical and immunological characterization, and molecular cloning of a cytoplasmic motor in a "lower" eukaryote, the ascomycete fungus neurospora crassa. this motor, termed nkin (for neurospora kinesin), exhibits several un ...19958589459
alterations in growth rate associated with a normally persisting circadian rhythm during spaceflight.biological clocks time many physiological parameters with periodicities close to 24 h; those which persist in the absence of environmental cues are circadian. an earlier shuttle experiment (sts-9) examined circadian pacemaker function and growth rate of neurospora crassa and demonstrated damped rhythm amplitudes, increased variability in period lengths and altered growth rates.19958588798
chromosome rearrangements that involve the nucleolus organizer region in neurospora.in approximately 3% of neurospora crassa rearrangements, part of a chromosome arm becomes attached to the nucleolus organizer region (nor) at one end of chromosome 2 (linkage group v). investigations with one inversion and nine translocations of this type are reported here. they appear genetically to be nonreciprocal and terminal. when a rearrangement is heterozygous, about one-third of viable progeny are segmental aneuploids with the translocated segment present in two copies, one in normal pos ...19958582636
binding affinity and functional significance of nit2 and nit4 binding sites in the promoter of the highly regulated nit-3 gene, which encodes nitrate reductase in neurospora crassa.in the filamentous fungus neurospora crassa, both the global-acting regulatory protein nit2 and the pathway-specific regulatory protein nit4 are required to turn on the expression of the nit-3 gene, which encodes nitrate reductase, the first enzyme in the nitrate assimilatory pathway. three nit2 binding sites and two nit4 binding sites have been identified in the 1.3-kb nit-3 promoter region via mobility shift and footprinting experiments with nit2-beta-galactosidase and nit4-beta-gactosidase fu ...19957592372
quantitative and qualitative aspects of spontaneous specific-locus mutation in the ad-3 region of heterokaryon 12 of neurospora crassa.the data from forward-mutation experiments to obtain specific-locus mutations at 2 closely linked loci in the adenine-3 (ad-3) region of heterokaryon 12 (h-12) of neurospora crassa have been tabulated to determine the frequency of spontaneous ad-3 mutations and to determine the percentages resulting from each of the 2 major genotypic classes: gene/point mutations and multilocus deletion mutations. gene/point mutations at the ad-3b locus (ad-3br) have been characterized to determine the percentag ...19957500991
inactivation of a single-2a phosphoprotein phosphatase is lethal in neurospora crassa.a pcr approach, employing the use of degenerate oligonucleotide mixtures, was used to isolate pph-1, a type-2a protein phosphatase (catalytic subunit)-encoding gene, from neurospora crassa. the isolated single copy gene is 1327 nucleotides in length, contains four putative introns and encodes a 310 amino-acid polypeptide. pph-1 is located between pdx-1 and col-4 on the right arm of n.crassa linkage group iv. pph-1 transcript levels are highest during the first hours of conidial germination. fail ...19958575020
dna methylation associated with repeat-induced point mutation in neurospora crassa.repeat-induced point mutation (rip) is a process that efficiently detects dna duplications prior to meiosis in neurospora crassa and peppers them with g:c to a:t mutations. cytosine methylation is typically associated with sequences affected by rip, and methylated cytosines are not limited to cpg dinucleotides. we generated and characterized a collection of methylated and unmethylated amrip alleles to investigate the connection(s) between dna methylation and mutations by rip. alleles of am harbo ...19957565710
cytochrome c oxidase in neurospora crassa contains myristic acid covalently linked to subunit 1.radiolabel from [3h]myristic acid was incorporated by neurospora crassa into the core catalytic subunit 1 of cytochrome c oxidase (ec 1.9.3.1), as indicated by immunoprecipitation. this modification of the subunit, which was specific for myristic acid, represents an uncommon type of myristoylation through an amide linkage at an internal lysine, rather than an n-terminal glycine. the [3h]myristate, which was chemically recovered from the radiolabeled subunit peptide, modified an invariant lys-324 ...19957567996
purification of neurospora crassa alkaline phosphatase without dnase activity for use in molecular biology.alkaline phosphatase, excreted by neurospora crassa preg (c) and purified to apparent homogeneity by 7.5% page, did not show dnaase activity and removed the terminal 5'-phosphate group from plasmid bluescript m13(+) linearized with ecori. the preg (c) strain may therefore replace other sources of alkaline phosphatase for use in dephosphorylating linearized plasmidial dna in cloning experiments.199524414902
crystallization and preliminary x-ray analysis of the nadp-specific glutamate dehydrogenase from neurospora crassa.the nadp-linked glutamate dehydrogenase from neurospora crassa has been crystallized by the hanging-drop method of vapour diffusion in the presence of 0.1 m glutamate. the crystals are trigonal and are in space group p3(1)21 with unit-cell dimensions of a = b = 196.6, c = 102.0 a and with a trimer in the asymmetric unit. a full structure determination of this enzyme will lead to an understanding of the molecular basis of inter-allelic complementation observed with hybrid hexamers of naturally oc ...199515299820
species-specific and mating type-specific dna regions adjacent to mating type idiomorphs in the genus neurospora.mating type idiomorphs control mating and subsequent sexual development in neurospora crassa and were previously shown to be well conserved in other neurospora species. the centromere-proximal flanks of the a and a idiomorphs, but not the distal flanks from representative heterothallic, pseudohomothallic, and homothallic neurospora species contain apparent species-specific and/or mating type-specific sequences adjacent to the well-conserved idiomorphs. the variable flank is bordered by regions t ...19958536961
mechanism of pyrithione-induced membrane depolarization in neurospora crassa.pyrithione is a general inhibitor of membrane transport in fungi and is widely used in antidandruff shampoos as an antifungal agent. an electrophysiological approach has been used to determine the mode of action of pyrithione on the plasma membrane of the model ascomycete, neurospora crassa. at ph 5.8, pyrithione induces a dramatic dose-dependent electrical depolarization of the membrane which is complete within 4 min, amounts to 110 mv at saturating pyrithione concentrations, and is half maxima ...19957574648
three-to-one segregation from reciprocal translocation quadrivalents in neurospora and its bearing on the interpretation of spore-abortion patterns in unordered asci.in neurospora, viable ascospores become black (b) when mature, whereas ascospores that are deficient for a chromosome segment are inviable and usually fail to blacken. the presence of a chromosome rearrangement can be recognized and the type of rearrangement can usually be inferred by visual inspection of asci. when a cross is heterozygous for a reciprocal translocation, asci with eight black ascospores (8b:0w) and asci with eight abortive unpigmented ("white" (w)) ascospores (0b:8w) are theoret ...19957672602
characterization of the promoter region of a cell-adhesion protein gene derived from the basidiomycete lentinus edodes.an analysis of the 2 kb nucleotide sequence including the 5'-flanking region of a cell-adhesion protein-encoding gene (mfba) isolated from the basidiomycete lentinus edodes revealed that the promoter region contains a tata box, a gc box, a caat box, a ct-rich sequence element, a tata box, two ct-rich sequences, and a caat box, in the order, from upstream to downstream. three major and three alternative transcriptional initiation sites were located 127, 129 and 131 nucleotides and 96, 193 and 197 ...19957649440
light and development regulate the expression of the albino-3 gene in neurospora crassa.the albino-3 gene of neurospora crassa codes for geranylgeranyl pyrophosphate synthase, an enzyme involved in the biosynthesis of carotenoids. the albino-3 locus encodes two overlapping transcription units that give rise to two mrnas of 2.2 kb (al-3(c)) and 1>6 kb (al-3(m)), with the promoter of the latter residing in the transcribed region of the former. the 1.6-kb transcript was transiently expressed in the mycelium after light induction, while the 2.2-kb mrna appeared in conidiating cultures ...19957649389
fourier transform infrared spectroscopy study of the secondary structure of the reconstituted neurospora crassa plasma membrane h(+)-atpase and of its membrane-associated proteolytic peptides.we reconstituted purified plasma membrane h(+)-atpase from neurospora crassa into soybean phospholipid vesicles (lipid/atpase ratio of 5:1 w/w). the proteoliposomes contained an active atpase, oriented inside-out. they were subjected to proteolysis by using pronase, proteinase k, trypsin, and carboxypeptidase y. fourier transform infrared attenuated total reflection spectroscopy indicates that the amount of protein remaining after hydrolysis and elimination of the extramembrane domain of atpase ...19957629067
a chimeric light-regulated amino acid transport system allows the isolation of blue light regulator (blr) mutants of neurospora crassa.we have developed a system for the isolation of neurospora crassa mutants that shows altered responses to blue light. to this end we have used the light-regulated promoter of the albino-3 gene fused to the neutral amino acid permease gene mtr. the product of the mtr gene is required for the uptake of neutral aliphatic and aromatic amino acids, as well as toxic analogs such as p-flurophenylalanine or 4-methyltryptophan. mtr trp-2-carrying cells were transformed with the al-3 promoter-mtr wild-typ ...19957604041
the chromosomal region which includes the recombinator cog in neurospora crassa is highly polymorphic.the st lawrence st74-or23-iva and lindegren y8743 strains of neurospora crassa have a different provenance from wild collections and dissimilar cog alleles; that in lindegren, cogla (previously designated cog+), is a more efficient recombinator than cogs74a and cogea (previously cog), the alleles in st lawrence and emerson a respectively. restriction fragment length polymorphisms (rflps) and sequence polymorphisms (sps) were used to map the difference between cogla and cogs74a to a region that e ...19958590467
analysis of the iron-sulfur clusters within the complex i (nadh:ubiquinone oxidoreductase) isolated from potato tuber mitochondria.the mitochondrial complex i (nadh:ubiquinone oxidoreductase) isolated from potato (solanum tuberosum) has been investigated for the presence of iron-sulfur clusters. epr spectroscopic analysis detected signals arising from clusters n1, n2, n3 and n4. quantitation of the content of iron and sulfur within the isolated complex i showed the preparation to contain 22.6 mol acid-labile sulfide and 30.4 mol iron/mol complex i. the iron-sulfur cluster composition of the plant complex i appears to be sim ...19957601133
a fungal actin-related protein involved in nuclear migration.the ro-4 mutant of the filamentous fungus neurospora crassa forms distinctive colonies in which hyphae grow into rope-like aggregates. this unusual morphology coincides with a defect in hyphal nuclear migration. the ro-4 gene was cloned from a cosmid library by complementation of the closely linked pab-2 gene. the deduced 380 amino acid protein is most similar to the vertebrate actin-related protein/centractin. the ro4 protein is not essential for cell viability, and new strains created by induc ...19957603438
redox imbalance at the start of each morphogenetic step of neurospora crassa conidiation.the conidiation process of neurospora crassa is characterized by three morphogenetic steps: hyphal adhesion, aerial hyphal formation, and production of conidia. total protein oxidation and specific enzyme oxidation coincided with an increased oxygen-dependent chemiluminescence and indicated the occurrence of a hyperoxidant state at the onset of all three morphogenetic steps. oxidation of nad(p)h and excretion of glutathione disulfide was detected at the start of hyphae adhesion. here we show tha ...19957786037
solubilization of neurospora crassa rodlet proteins and identification of the predominant protein as the proteolytically processed eas (ccg-2) gene product.proteins from conidial rodlet preparations of neurospora crassa were solubilized in trifluoroacetic acid. sodium dodecyl sulfate-polyacrylamide gel electrophoresis of solubilized rodlets revealed a predominant protein of approximately 7 kda. this protein was absent from preparations of n. crassa cultures carrying the eas mutation. the protein was purified by reverse-phase high-performance liquid chromatography and the n-terminal amino acid sequence of the purified protein was found to be identic ...19957614378
resistance to azole drugs in neurospora crassa.neurospora crassa was susceptible to azole drugs: ketoconazole (mic 1 microgram/ml), fluconazole (mic 5 micrograms/ml), and sch39304 (mic 5 micrograms/ml). mutants of n. crassa resistant to ketoconazole were selected and genetically characterized. the seven characterized resistance mutations represented at least four genetic loci. some mutants, but not all, were also resistant to fluconazole and to sch39304.19957614377
[analysis of light signal transmission through the phosphorylation of proteins in neurospora crassa]. 19957610259
translocation of apocytochrome c across the outer membrane of mitochondria.apocytochrome c follows a unique pathway into mitochondria. import does not require the general protein translocation machinery, protease-sensitive components of the outer membrane, or a membrane potential across the inner membrane. we investigated the membrane binding and translocation steps of the import reaction using purified outer membrane vesicles (omv) from neurospora crassa mitochondria. omv specifically bound, but did not import apocytochrome c. if, however, specific antibodies were enc ...19957759479
reconstitution of the neurospora crassa plasma membrane h(+)-adenosine triphosphatase.the purified h(+)-atpase of the neurospora crassa plasma membrane has been reconstituted by a gel filtration method into lipidic vesicles using sodium deoxycholate as the detergent. reconstitution was performed for lipid/atpase ratios ranging from 1000:1 to 5:1 (w/w). whatever the lipid/atpase ratio, the atpase molecules completely associate with the lipid vesicles. the atpase specific activity is identical for all proteoliposomes regardless of the lipid/atpase ratio, but the h+ transport decrea ...19957794959
developmental and photoregulation of al-1 and al-2, structural genes for two enzymes essential for carotenoid biosynthesis in neurospora.the levels of al-1 and al-2 transcripts change dramatically in response to light and development during the formation of neurospora crassa asexual spores (conidia). al-1 and al-2 mrnas accumulate throughout conidiation irrespective of lighting conditions initially at low levels. as conidiation proceeds, two increases in albino message accumulation are observed. this developmentally induced photoindependent message accumulation was not observed in neurospora mutants blocked at relevant stages of ...19957750660
2-d structure of the neurospora crassa plasma membrane atpase as determined by electron cryomicroscopy.large, well-ordered 2-d crystals of the dodecylmaltoside complex of the neurospora crassa plasma membrane h(+)-atpase grow rapidly on the surface of a polyethylene glycol-containing mixture similar to that originally developed for growing 3-d crystals of this integral membrane transport protein. negative stain electron microscopy of the crystals shows that many are single layers. cryoelectron microscopy of unstained specimens indicates that the crystals have a p6 layer group with unit cell dimen ...19957743992
cyclophilin 20 is involved in mitochondrial protein folding in cooperation with molecular chaperones hsp70 and hsp60.we studied the role of mitochondrial cyclophilin 20 (cyp20), a peptidyl-prolyl cis-trans isomerase, in preprotein translocation across the mitochondrial membranes and protein folding inside the organelle. the inhibitory drug cyclosporin a did not impair membrane translocation of preproteins, but it delayed the folding of an imported protein in wild-type mitochondria. similarly, neurospora crassa mitochondria lacking cyp20 efficiently imported preproteins into the matrix, but folding of an import ...19957739545
light induction of the clock-controlled gene ccg-1 is not transduced through the circadian clock in neurospora crassa.ambient light and the circadian clock have been shown to be capable of acting either independently or in an interrelated fashion to regulate the expression of conidiation in the ascomycete fungus neurospora crassa. recently several molecular correlates of the circadian clock have been identified in the form of the morning-specific clock-controlled genes ccg-1 and ccg-2. in this paper we report studies on the regulation of ccg-1, an abundantly expressed gene displaying complex regulation. consist ...19957753024
the sulfur controller-2 negative regulatory gene of neurospora crassa encodes a protein with beta-transducin repeats.the sulfur regulatory system of neurospora crassa is composed of a set of structural genes involved in sulfur catabolism controlled by a genetically defined set of trans-acting regulatory genes. these sulfur regulatory genes include cys-3+, which encodes a basic region-leucine zipper transcriptional activator, and the negative regulatory gene scon-2+. we report here that the scon-2+ gene encodes a polypeptide of 650 amino acids belonging to the expanding beta-transducin family of eukaryotic regu ...19957724564
guest: a 98 bp inverted repeat transposable element in neurospora crassa.the region immediately 3' of histidine-3 has been cloned and sequenced from two laboratory strains of the ascomycete fungus neurospora crassa; st lawrence 74a and lindegren, which have different derivations from wild collections. amongst the differences distinguishing these sequences are insertions ranging in size from 20 to 101 bp present only in st lawrence. the largest of these is flanked by a 3 bp direct repeat, has terminal inverted repeats (tir) and shares features with several known trans ...19957715596
some property of the nucleus determines the competence of neurospora crassa for transformation.in neurospora, transformation of spheroplasts is quite efficient and usually occurs with the transforming dna integrated at ectopic sites in the chromosome. however, only a small fraction of the spheroplasts is actually competent for transformation. to distinguish whether the limitation to competence is at the level of the plasma membrane or at the level of the nucleus, we performed experiments in which heterocaryotic spheroplasts were required to integrate two different plasmids in one transfor ...19957789759
cloning, nucleotide sequence, and expression of tef-1, the gene encoding translation elongation factor 1 alpha (ef-1 alpha) of neurospora crassa.the tef-1 gene encoding translation elongation factor 1 alpha was cloned from the ascomycete fungus neurospora crassa. the sequences of genomic dna and cdna clones showed that the tef-1 gene contained one orf of 1380 bp length that is interrupted by three short introns. the deduced polypeptide contained 460 amino acid residues, and the sequence had a high similarity with those of ef-1 alpha polypeptides from other species. the level of tef-1 mrna was low in conidia but high in growing cells. whe ...19957605676
'sheltered disruption' of neurospora crassa mom22, an essential component of the mitochondrial protein import complex.mom22 is a component of the protein import complex of the mitochondrial outer membrane of neurospora crassa. using the newly developed procedure of 'sheltered disruption', we created a heterokaryotic strain harboring two nuclei, one with a null allele of the mom-22 gene and the other with a wild-type allele. homokaryons bearing the mom-22 disruption could not be isolated, suggesting that mom-22 is an essential gene. the mutant nucleus can be forced to predominate in the heterokaryon through the ...19957720701
close correlation between heat shock response and cytotoxicity in neurospora crassa treated with aliphatic alcohols and phenols.in neurospora crassa the aliphatic alcohols methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, isobutanol, ethylene glycol, glycerol, and allyl alcohol and the phenolic compounds phenol, hydroquinone, resorcinol, pyrogallol, phloroglucinol, sodium salicylate, and acetylsalicylic acid were analyzed with respect to their capacities to induce heat shock proteins (hsp) and to inhibit protein synthesis. both the alcohols and phenols showed the greatest levels of hsp induction at concent ...199516534981
inactivation of genes encoding subunits of the peripheral and membrane arms of neurospora mitochondrial complex i and effects on enzyme assembly.we have isolated and characterized the nuclear genes encoding the 12.3-kd subunit of the membrane arm and the 29.9-kd subunit of the peripheral arm of complex i from neurospora crassa. the former gene was known to be located in linkage group i and the latter is now assigned to linkage group iv of the fungal genome. the genes were separately transformed into different n. crassa strains and transformants with duplicated dna sequences were isolated. selected transformants were then mated with other ...19957768434
genetic nomenclature guide. neurospora crassa. 19957660461
disruption of the gene encoding the 78-kilodalton subunit of the peripheral arm of complex i in neurospora crassa by repeat induced point mutation (rip).we have used the procedure of sheltered rip to generate mutants of the 78-kda protein of the peripheral arm of neurospora crassa complex i. the nuclei containing the mutations were initially isolated as one component of a heterokaryon but subsequent analysis showed that nuclei containing null alleles of the gene could be propagated as homokaryons. this demonstrates that the gene does not serve an essential function. sequence analysis of one allele shows that 61 transition mutations were created ...19957614557
the fungal h(+)-atpase from neurospora crassa reconstituted with fusicoccin receptors senses fusicoccin signal.fusicoccin affects several physiological processes regulated by the plasma membrane h(+)-atpase in higher plants while other organisms having p-type h(+)-atpases (e.g., fungi) are fusicoccin-insensitive. we have previously shown that fusicoccin binding to its receptor is necessary for h(+)-atpase stimulation and have achieved the functional reconstitution into liposomes of fusicoccin receptors and the h(+)-atpase from maize. in this paper we show that fusicoccin sensitivity can be conferred on t ...19957878025
folylpolyglutamate synthesis in neurospora crassa: transformation of polyglutamate-deficient mutants.the methionine auxotrophy of neurospora crassa met-6 and mac mutants is related to an inability to synthesize long-chained folylpolyglutamates. both of these lesions affect folylpolyglutamate synthetase activity, but it is not clear whether these mutations occur in different genes or in functional domains of the same gene. to address this question, copies of the met-6+ gene have been introduced into both mutants using plasmid and cosmid vectors. transformation to prototrophy was achieved in both ...19957766162
the vacuolar atpase: sulfite stabilization and the mechanism of nitrate inactivation.using vacuolar membranes from neurospora crassa, we observed that sulfite prevented the loss of vacuolar atpase activity that otherwise occurred during 36 h at room temperature. sulfite neither activated nor changed the kinetic behavior of the enzyme. further, in the presence of sulfite, the vacuolar atpase was not inhibited by nitrate. we tested the hypothesis that sulfite acts as a reducing agent to stabilize the enzyme, while nitrate acts as an oxidizing agent, inhibiting the enzyme by promot ...19957829484
observation of a secondary tritium isotope effect in the chorismate synthase reaction.chorismate synthase, the seventh enzyme on the shikimate pathway, catalyzes the formation of chorismate from 5-enolpyruvylshikimate 3-phosphate (epsp). this reaction involves the loss of phosphate from c(3) and hydrogen from the c(6) pro-r position of epsp. in order to probe the mechanism of this reaction, [3-(3)h, 14c]epsp has been synthesized and a secondary v/k tritium kinetic isotope measured for the reaction catalyzed by neurospora crassa chorismate synthase. a small but significant value o ...19957819217
purification of deoxyhypusine synthase from neurospora crassa to homogeneity by substrate elution affinity chromatography.deoxyhypusine synthase is an nad(+)-dependent enzyme that catalyzes the formation of deoxyhypusine residue on the eif-5a precursor by using spermidine as the substrate. deoxyhypusine synthase bound tightly to 1,12-diaminododecane-agarose and could be eluted selectively by spermidine. this finding enabled us to develop a simple two-column procedure to purify deoxyhypusine synthase from neurospora crassa to apparent homogeneity. the purified enzyme had a specific activity of 130,000 units/mg of pr ...19957814398
a stringent role of f-actin in the ascogonial differentiation of neurospora crassa fluffy mutant.cytochalasin d, an inhibitor of actin polymerization, interferes with ascogonial differentiation in the fertile fluffy mutant of neurospora crassa. as the total level of actin and its mrna remain unchanged, this suggests that it is in its microfilamentous form (f-form) that actin is stringently required for female differentiation.19958768259
ubiquinol-cytochrome-c oxidoreductase of neurospora crassa. 19958592472
biosynthesis of pteridines in neurospora crassa, phycomyces blakesleeanus and euglena gracilis: detection and characterization of biosynthetic enzymes.occurrence, biosynthesis and some functions of tetrahydrobiopterin (h4biopterin) in animals are well known. the biochemistry of h4biopterin in other organisms than animals was hitherto not widely investigated. recently h4biopterin was found in the phytoflagellate euglena gracilis, in the zygomycete phycomyces blakesleeanus and in the ascomycete neurospora crassa. in euglena, neurospora and phycomyces the enzymatic activities of gtp cyclohydrolase i, 6-pyruvoyl tetrahydropterin synthase and sepia ...19957899493
the hsp70 gene family of neurospora crassa: cloning, sequence analysis, expression, and genetic mapping of the major stress-inducible member.the gene encoding the major heat shock-inducible member of the hsp70 family of neurospora crassa was cloned and characterized. the 5' nontranscribed region shows the presence of consensus sequence motifs resembling the classical heat shock elements found in many heat shock-responsive eukaryotic promoters, as well as metal-responsive-element sequences. the coding region of the gene contains four introns with boundaries and internal consensus motifs typical of genes of filamentous fungi. none of t ...19957798134
isolation of dihydrofolate and folylpolyglutamate synthetase activities from neurospora.the possible association of dihydrofolate synthetase (dhfs) and folylpolyglutamate synthetase (fpgs) in neurospora crassa (fgsc 853, wild type) has been examined using mycelial extracts prepared and fractionated in the presence of protease inhibitors. dhfs and fpgs were assayed by following the incorporation of labelled glutamate into dihydrofolate and methylenetetrahydrofolate polyglutamate, respectively. both of these activities were predominately cytosolic in mycelia that were harvested 24 hr ...19957772303
spontaneous mutation during the sexual cycle of neurospora crassa.the dna sequences of 42 spontaneous mutations of the mtr gene in neurospora crassa have been determined. the mutants were selected among sexual spores to represent mutations arising in the sexual cycle. three sexual-cycle-specific mutational classes are described: hotspot mutants, spontaneous repeat-induced point mutation (rips) and mutations occurring during a mutagenic phase of the sexual cycle. together, these three sexual-cycle-specific mutational classes account for 50% of the mutations in ...19957705619
purification and characterization of mitochondrial processing peptidase of neurospora crassa. 19957674958
the effects of temperature change on the circadian clock of neurospora.the phase resetting of the circadian oscillatory system by pulses of increased temperature (zeitgebers) and the temperature compensation of its period length during longer exposures are major features of the system, but are not well understood in molecular terms. in neurospora crassa, the effects of pulses of increased temperature on the circadian rhythm of conidiation were determined and possible inputs to the oscillatory system tested, including changes in cyclic 3',5'-adenosine monophosphate ...19957656690
nuclear actin and rna export in conidial cells of neurospora crassa. 19957542118
genetic and epigenetic inactivation of repetitive sequences in neurospora crassa: rip, dna methylation, and quelling. 19957493491
a tyrosyl-trna synthetase recognizes a conserved trna-like structural motif in the group i intron catalytic core.the neurospora crassa mitochondrial (mt) tyrosyl-trna synthetase (cyt-18 protein) functions in splicing group i introns, in addition to aminoacylating trna(tyr). here, we compared the cyt-18 binding sites in the n. crassa mt lsu and nd1 introns with that in n. crassa mt trna(tyr) by constructing three-dimensional models based on chemical modification and rna footprinting data. remarkably, superimposition of the cyt-18 binding sites in the model structures revealed an extended three-dimensional o ...19968978617
a dominant negative effect of eth-1r, a mutant allele of the neurospora crassa s-adenosylmethionine synthetase-encoding gene conferring resistance to the methionine toxic analogue ethionine.eth-1r, a thermosensitive allele of the neurospora crassa s-adenosylmethionine (adomet) synthetase gene that confers ethionine resistance, has been cloned and sequenced. replacement of an aspartic amino acid residue (d48-->n48), perfectly conserved in prokaryotic, fungal and higher eukaryotic adomet synthetases, was found responsible for both thermosensitivity and ethionine resistance conferred by eth-1r. gene fusion constructs, designed to overexpress eth-1r in vivo, render transformant cells r ...19968978034
role of cytosolic serine hydroxymethyltransferase in one-carbon metabolism in neurospora crassa.conidiospores of wild type and two mutant strains of neurospora crassa were grown on [3-13c]serine, [2-13c]glycine, or [13c]formate. acid extracts of the mycelia were analyzed by 13c nmr for incorporation of the 13c label into choline, serine, and adenine. the goal was to elucidate the function of cytosolic serine hydroxymethyltransferase by comparison of a mutant strain lacking this activity and requiring formate for optimal growth (for mutant strain) to the wild-type strain and the ser3 strain ...19968914930
circadian clock-controlled genes isolated from neurospora crassa are late night- to early morning-specific.an endogenous circadian biological clock controls the temporal aspects of life in most organisms, including rhythmic control of genes involved in clock output pathways. in the fungus neurospora crassa, one pathway known to be under control of the clock is asexual spore (conidia) development. to understand more fully the processes that are regulated by the n. crassa circadian clock, systematic screens were carried out for genes that oscillate at the transcriptional level. time-of-day-specific cdn ...19968917550
metabolism of ricinoleate by neurospora crassa.neurospora crassa is a potential expression system for evaluating fatty-acid-modifying genes from plants producing uncommon fatty acids. one such gene encodes the hydroxylase that converts oleate to ricinoleate, a fatty acid with important industrial uses. to develop this expression system, it is critical to evaluate the metabolism and physiological effects of the expected novel fatty acid(s). we therefore examined effects of ricinoleate on lipid biosynthesis and growth of n. crassa. ricinoleate ...19968987727
acetylglutamate synthase from neurospora crassa: structure and regulation of expression.a dna clone which complemented an arg-14 mutation of neurospora crassa was isolated by sib selection from a cosmid library (pmocosx). southern and restriction-fragment-length polymorphism (rflp) analysis confirmed that the cloned dna contained the arg-14 gene. the arg-14 gene was identified as the structural gene for acetylglutamate synthase by immunodepletion of enzyme activity with antibodies prepared against an arg-14 fusion protein and by the thermal instability of acetylglutamate synthase i ...19968939437
loss of growth polarity and mislocalization of septa in a neurospora mutant altered in the regulatory subunit of camp-dependent protein kinase.in filamentous fungi, growth polarity (i.e. hyphal extension) and formation of septa require polarized deposition of new cell wall material. to explore this process, we analyzed a conditional neurospora crassa mutant, mcb, which showed a complete loss of growth polarity when incubated at the restrictive temperature. cloning and dna sequence analysis of the mcb gene revealed that it encodes a regulatory subunit of camp-dependent protein kinase (pka). unexpectedly, the mcb mutant still formed sept ...19968918454
secondary and tertiary structure changes of reconstituted p-glycoprotein. a fourier transform attenuated total reflection infrared spectroscopy analysis.the structure of purified p-glycoprotein functionally reconstituted into liposomes was investigated by attenuated total reflection fourier transform infrared spectroscopy. a quantitative evaluation of the secondary structure and a kinetic of 2h/h exchange of the p-glycoprotein were performed both in the presence and in the absence of mgatp, mgatp-verapamil, and mgadp. this approach was previously shown to be a useful tool to detect tertiary structure changes resulting from the interaction betwee ...19968798727
intracellular phosphate--water oxygen exchange measured by mass spectrometry.to study, in vivo, potential p(i)-water oxygen exchange catalyzed by each of two high-affinity p(i) symporters of neurospora crassa, we have developed methods for the purification of p(i) from whole-cell extracts and the subsequent derivatization of p(i) for analysis by gc-ms. we have also modified a published procedure for the preparation of 18o-p(i). however, the high background rate of transport-independent oxygen exchange, determined by monitoring the appearance of 18o-p(i) in cells incubate ...19968921158
escape from het-6 incompatibility in neurospora crassa partial diploids involves preferential deletion within the ectopic segment.self-incompatible het-6or/het-6pa partial diploids of neurospora crassa were selected from a cross involving the translocation strain, t(iil-->iiir)ar18, and a normal sequence strain. about 25% of the partial diploids exhibited a marked increase in growth rate after 2 weeks, indicating that "escape" from het-6 incompatibility had occurred. near isogenic tester strains with different alleles (het-6or and het-6pa) were constructed and used to determine that 80 of 96 escape strains tested were het- ...19968889517
mild rip-an alternative method for in vivo mutagenesis of the albino-3 gene in neurospora crassa.we have used a biological phenomenon that occurs in neurospora crassa, termed repeat-induced point mutation (rip), to create partially functional mutant alleles of the albino-3 (al-3) gene encoding geranylgeranyl pyrophosphate synthase, an enzyme involved in the biosynthesis of carotenoids and diverse prenylated compounds. a total of 70 rip-induced al-3 mutants were identified by their pale albino phenotype, resulting from inactivation of carotenoid bio-synthesis. nucleotide sequence analysis of ...19968879235
bli-4, a gene that is rapidly induced by blue light, encodes a novel mitochondrial, short-chain alcohol dehydrogenase-like protein in neurospora crassa.blue light plays an important role in developmental control throughout nature. the bli-4 gene of neurospora crassa, together with bli-3, al-1 and al-2, is rapidly inducible by blue light. induction leads to a ninety-fold increase in transcription rate over the dark control level, and the gene therefore appears to be of prime importance in the blue-light induction pathway of n. crassa. we describe the sequencing and analysis of bli-4 and the 38 kda protein it encodes. we show that the protein is ...19968842141
translocation of neurospora crassa transcription factor nuc-1 into the nucleus is induced by phosphorus limitation.nuc-1, a basic helix-loop-helix zipper protein, activates the expression of several genes involved in phosphorus acquisition in neurospora crassa. in the present study we investigated whether posttranscriptional mechanisms control the activity of nuc-1. the nuc-1 level was higher (up to fivefold) in wild-type cells grown at low external phosphate concentration and in mutant strains expressing the phosphorus acquisition genes constitutively than in a wild-type strain grown at high external phosph ...19968953266
light-induced phase shifting of the circadian conidiation rhythm is inhibited by calmodulin antagonists in neurospora crassa.the effects of calmodulin antagonists and inhibitors of protein kinases and phosphatases on light-induced phase shifting were investigated in neurospora crassa. calmodulin antagonists, namely, trifluoperazine, chlorpromazine, and w-7, almost completely inhibited the light-induced phase shifting without having any effect on the circadian clock itself. chlorpromazine was less effective in inhibiting the light-induced phase shifting than trifluoperazine. w-5, a dechlorinated analogue of w-7, failed ...19968872595
downstream 28s gene sequences on the rna template affect the choice of primer and the accuracy of initiation by the r2 reverse transcriptase.r2 non-long terminal repeat retrotransposable elements insert at a unique site in the 28s rrna genes of insects. the protein encoded by the single open reading frame of r2 is capable of conducting the initial steps of its integration in vitro. the protein nicks the noncoding strand of the 28s target dna (the strand which serves as a template for rna synthesis) and uses the 3' hydroxyl group exposed by this nick to prime reverse transcription of the r2 rna template. this target-primed reverse tra ...19968756630
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