| functional roles and subsite locations of leu177, trp178 and asn182 of aspergillus awamori glucoamylase determined by site-directed mutagenesis. | fungal glucoamylases contain four conserved regions. one region from the aspergillus niger enzyme contains three key carboxylic acid residues, the general acid catalytic group, glu179, along with asp176 and glu180. three site-directed mutations, leu177-->his, trp178-->arg and asn182-->ala, were constructed near these acidic groups to reveal the function of other conserved residues in this region. leu177 and trp178 are strictly conserved among fungal glucoamylases, while an amide, predominantly a ... | 1993 | 8433972 |
| molecular analysis and overexpression of the gene encoding endothiapepsin, an aspartic protease from cryphonectria parasitica. | the gene, epn-1, encoding endothiapepsin (epn), an aspartic protease (aspp) synthesized and secreted by the ascomycete fungus responsible for chestnut blight, cryphonectria (endothia) parasitica, was identified and characterized. inspection of the nucleotide and deduced amino acid (aa) sequences revealed perfect agreement with the experimentally derived 330-aa sequence of mature epn [barkholt, eur. j. biochem. 167 (1987) 327-338] and an additional 89 aa of putative preprosequence. of the nine fu ... | 1993 | 8462868 |
| identification and elimination by site-directed mutagenesis of thermolabile aspartyl bonds in aspergillus awamori glucoamylase. | both native aspergillus niger glucoamylase and wild-type aspergillus awamori glucoamylase expressed in saccharomyces cerevisiae, which have identical primary structures, undergo hydrolysis at aspartyl bonds at low ph values and elevated temperatures. in native a.niger enzyme the asp126-gly127 bond was preferentially cleaved at ph 3.5, while at ph 4.5 cleavage of the asp257-pro258 and asp293-gly294 bonds was dominant. in wild-type a.awamori glucoamylase, cleavage of the latter was dominant at bot ... | 1995 | 8532682 |
| development of yeast strains for the efficient utilisation of starch: evaluation of constructs that express alpha-amylase and glucoamylase separately or as bifunctional fusion proteins. | eight constructions involving the bacillus subtilis alpha-amylase gene (amye), a mouse pancreatic alpha-amylase cdna (amy2) and an aspergillus awamori glucoamylase cdna (glaa) were prepared: three fusion genes, involving one alpha-amylase and the glucoamylase, two double-cassette plasmids (expressing one or other alpha-amylase and the glucoamylase) and three single-cassette plasmids, expressing the individual coding sequences. following transformation of each plasmid into saccharomyces cerevisia ... | 1995 | 8590658 |
| functional and structural roles of the highly conserved trp120 loop region of glucoamylase from aspergillus awamori. | the functional role of a loop region, highly conserved among glucoamylase and other starch hydrolases which also includes the essential trp120 of aspergillus awamori, is investigated. residues 121-125 of a. awamori glucoamylase were singly substituted, and their individual effects on catalytic activity and thermal stability were determined. the arg122-->tyr mutation displayed opposing effects for shorter and longer maltooligosaccharide substrates, k(m) decreasing for shorter substrates but incre ... | 1996 | 8608145 |
| identification and cloning of a mobile transposon from aspergillus niger var. awamori. | aspergillus niger var. awamori contains multiple copies of a transposable element, vader. this element was detected as a 437-bp insertion in four independently isolated spontaneous mutants of the niad (nitrate reductase) gene. the vader element is present in approximately 15 copies in both a. niger var. awamori and a. niger. a single copy of vader was detected from only one of the two laboratory strains of a. nidulans which were also examined. insertion of the vader element into the niad gene of ... | 1996 | 8625427 |
| crystallographic complexes of glucoamylase with maltooligosaccharide analogs: relationship of stereochemical distortions at the nonreducing end to the catalytic mechanism. | crystal structures at ph 4 of complexes of glucoamylase from aspergillus awamori var. x100 with the pseudotetrasaccharides d-gluco-dihydroacarbose and acarbose have been refined to r-factors of 0.147 and 0.131 against data to 1.7- and 2.0-a resolution, respectively. the two inhibitors bind in nearly identical manners, each exhibiting a dual binding mode with respect to the location of the last sugar residues. the reduced affinity of d-gluco-dihydroacarbose (k1 = 10(-8) m) relative to acarbose (k ... | 1996 | 8679589 |
| mutational modulation of substrate bond-type specificity and thermostability of glucoamylase from aspergillus awamori by replacement with short homologue active site sequences and thiol/disulfide engineering. | rational protein engineering based on three-dimensional structure, sequence alignment, and previous mutational analysis served to increase thermostability and modulate bond-type specificity in glucoamylase from aspergillus awamori. the single free cysteine, cys320, became disulfide bonded in the ala246 --> cys mutant, thus enhancing t50 by 4 degrees c to 73 degrees c. compared to wild-type, ala246 --> cys was roughly twice as active at 66 degrees c, but half as active at 45 degrees c. the altern ... | 1996 | 8679632 |
| studies on glucoamylase produced from aspergillus awamori (nrrl-3112) and their effect on saccharification of potato starch. | efficiency of an enzymatic starch saccharification process depends not only on the activity of the glucoamylase but also its purity. about 96.82% unwanted proteins present in 2-day old culture broth of a. awamori nrrl-3112 (grown in mygp medium) were separated by precipitation with ammonium sulphate which was followed by dialysis. more than 80% activity of the glucoamylase was recovered. three protein fractions (a, b, c) were identified using gel permeation chromatography. fractions a and b show ... | 1995 | 8714078 |
| arabinoxylan-degrading enzyme system of the fungus aspergillus awamori: purification and properties of an alpha-l-arabinofuranosidase. | an alpha-l-arabinofuranosidase produced by the fungus aspergillus awamori had a molecular mass of approximately 64 kda on sodium dodecyl sulphate/polyacrylamide gel electrophoresis (sds-page) and was optimally active at ph 4.6 and 50 degrees c. the enzyme, which chromatographed as a single component on sds-page, appeared to consist of two isoenzymes of pi 3.6 and 3.2. acting in isolation, the alpha-l-arabinofuranosidase had only a very limited capacity to release l-arabinose (less than 11%) dire ... | 1996 | 8737575 |
| deletion analysis of the starch-binding domain of aspergillus glucoamylase. | the large form of glucoamylase (gai) from aspergillus awamori (ec 3.2.1.3) binds strongly to native granular starch, whereas a truncated form (gaii) which lacks 103 c-terminal residues, does not. this c-terminal region, conserved among fungal glucoamylases and other starch-degrading enzymes, is part of an independent starch-binding domain (sbd). to investigate the sbd boundaries and the function of conserved residues in two putative substrate-binding sites, five gluco-amylase mutants were constr ... | 1995 | 8771186 |
| analysis of heterologous protein production in defined recombinant aspergillus awamori strains. | a study was carried out to obtain more insight into the parameters that determine the secretion of heterologous proteins from filamentous fungi. a strategy was chosen in which the mrna levels and protein levels of a number of heterologous genes of different origins were compared. all genes were under control of the aspergillus awamori 1,4-beta-endoxylanase a (exla) expression signals and were integrated in a single copy at the a. awamori pyrg locus. a northern (rna) analysis showed that large di ... | 1996 | 8787393 |
| kinetics of mrna and protein synthesis of genes controlled by the 1,4-beta-endoxylanase a promoter in controlled fermentations of aspergillus awamori. | in this study, induction and repression kinetics of the expression of the aspergillus awamori 1,4-beta-endoxylanase a (exla) gene under defined physiological conditions was analyzed at the mrna and the protein levels. induction was analyzed by pulsing d-xylose to a sucrose-limited continuous culture of an a. awamori 1,4-beta-endoxylanase a (exla)-overproducing strain. directly after the d-xylose pulse, exia mrna was synthesized, and it reached a constant maximal level after 45 to 60 min. this le ... | 1996 | 8837419 |
| effect of replacing helical glycine residues with alanines on reversible and irreversible stability and production of aspergillus awamori glucoamylase. | to decrease irreversible thermoinactivation of aspergillus awamori glucoamylase, five gly residues causing helix flexibility were replaced with ala residues. mutation of gly57 did not affect thermostability. mutation of gly137 doubled it at phs 3.5 and 4.5 but barely changed it at ph 5.5. the gly139-->ala mutation did not change thermostability at ph 3.5, improved it at ph 4.5 and worsened it at ph 5.5. the gly 137/gly139-->ala/ala mutation gave 1.5-2-fold increased thermostabilities at phs 3.5- ... | 1996 | 8862550 |
| stereochemical course of hydrolysis catalyzed by arabinofuranosyl hydrolases. | the stereochemical course of hydrolysis catalyzed by various enzymes acting on arabinofuranosyl linkages has been determined. 1h-nmr analysis of the action of endo-(1-->5)-alpha-l-arabinanases from aspergillus niger and aspergillus aculeatus showed that both hydrolyze linear arabinan with inversion of configuration, and may therefore act via a single displacement mechanism. this is consistent with the a. niger enzyme's classification in glycosyl hydrolase family 43. the catalytic mechanisms of a ... | 1996 | 8946944 |
| identification of enzyme-substrate and enzyme-product complexes in the catalytic mechanism of glucoamylase from aspergillus awamori. | intermediates in the catalytic mechanism of aspergillus awamori glucoamylase (ga) were identified by studying pre-steady-state and steady-state kinetics of the wild-type ga/maltose and trp120 -->phe ga/maltotriose reactions in h2o and d2o. pre-steady-state fluorescence signal analysis was carried out to ascertain the relative intrinsic fluorescence of the enzyme intermediates. a three-step minimal pathway for oligosaccharide hydrolysis represented by e + gx (k1) reversible (k-1) egx (k2)reversib ... | 1996 | 8952477 |
| expression of human recombinant beta 2-microglobulin by aspergillus nidulans and its activity. | the light chain of hla class i protein (beta 2m) has been expressed in aspergillus nidulans. the cdna of beta 2m was modified using the polymerase chain reaction to include overlapping extensions for its subsequent fusion into an aspergillus vector. this fusion resulted in beta 2m cdna being flanked by the aspergillus awamori glucoamylase promoter and the aspergillus niger glucoamylase terminator. expression of beta 2m was induced by the addition of starch to the culture medium. in preliminary m ... | 1996 | 8960907 |
| efficient expression of a phanerochaete chrysosporium manganese peroxidase gene in aspergillus oryzae. | a manganese peroxidase gene (mnp1) from phanerochaete chrysosporium was efficiently expressed in aspergillus oryzae. expression was achieved by fusing the mature cdna of mnp1 with the a. oryzae taka amylase promoter and secretion signal. the 3' untranslated region of the glucoamylase gene of aspergillus awamori provided the terminator. the recombinant protein (rmnp) was secreted in an active form, permitting rapid detection and purification. physical and kinetic properties of rmnp were similar t ... | 1996 | 8975615 |
| the effect of pre- and pro-sequences and multicopy integration on heterologous expression of the fusarium solani pisi cutinase gene in aspergillus awamori. | a synthetic derivative of the cutinase cdna of fusarium solani pisi was expressed in aspergillus awamori using the a. awamori endoxylanase ii (exla) promoter and terminator. the influence of the origin of the pre-sequence and the presence of a pro-sequence on the efficiency of extracellular cutinase production was analysed in single-copy transformants containing an expression cassette integrated at the pyrg locus. transformants containing a construct encoding a direct, inframe fusion of the xyla ... | 1996 | 8987467 |
| an expression system based on the promoter region of the aspergillus awamori 1,4-beta-endoxylanase a gene. | a new, highly inducible fungal promoter derived from the aspergillus awamori 1,4-beta-endoxylanase a (exla) gene is described. induction analysis, carried out with the wild-type strain in shake flasks, showed that exla expression in regulated at the transcriptional level. using a beta-glucuronidase (uida) reporter strategy, d-xylose was shown to be an efficient inducer of the exla promoter, whereas sucrose or maltodextrin were not. upon d-xylose induction, the exla promoter was threefold more ef ... | 1996 | 8987532 |
| the transposable element tan1 of aspergillus niger var. awamori, a new member of the fot1 family. | aspergillus niger var. awamori has transposable elements that we refer to as vader and tan1 (transposon a. niger). vader was identified by screening unstable nitrate reductase (niad) mutants for insertions. four of the isolated niad mutants were shown to contain a small insertion element. this 437 bp insertion element, vader, is flanked by 44 bp inverted repeats (ir) and is present in approximately 15 copies in the genomes of two a. niger strains examined. a synthetic 44 bp oligomer of the inver ... | 1996 | 9003286 |
| glucoamylase gene fusions alleviate limitations for protein production in aspergillus awamori at the transcriptional and (post) translational levels. | in this study we have analyzed the effects of a glucoamylase gene fusion on the mrna levels and protein levels for the human interleukin-6 gene (hil6) and the guar alpha-galactosidase gene (agla). previously it was shown that production of nonfused alpha-galactosidase and hil-6 in aspergillus awamori was limited at transcriptional and (post)translational levels, respectively (r. j. gouka, p. j. punt, j. g. m. hessing, and c. a. m. j. j. van den hondel, appl. environ. microbiol. 62:1951-1957, 199 ... | 1997 | 9023927 |
| efficient production of secreted proteins by aspergillus: progress, limitations and prospects. | filamentous fungi are widely used for the production of homologous and heterologous proteins but, compared to homologous proteins, the levels of production of heterologous proteins are usually low. during the last 5 years, the levels of production of heterologous proteins have been drastically improved by fusing the corresponding gene to the 3' end of a homologous gene, encoding a well-secreted protein such as glucoamylase. nevertheless, little research has been carried out to determine the limi ... | 1997 | 9035405 |
| glucoamylase mutants in the conserved active-site segment trp170-tyr175 located at a distance from the site of catalysis. | to mimic the structure of the 1.8-fold more active (k(cat)) rhizopus oryzae glucoamylase (ga), aspergillus niger ga was subjected to site-directed mutagenesis in the trp170-tyr175 segment of the third of the six well-conserved alpha-->alpha connecting loops of the catalytic (alpha/alpha)6-barrel. while the trp170-->phe, gln172-->asn and tyr175-->phe mutants showed an up to 1.7-fold increased k(cat) and gly174-->cys ga and approximately 2-fold reduced k(cat) towards maltotriose and longer substra ... | 1997 | 9051738 |
| overexpression and characterization of aspergillus awamori wild-type and mutant glucoamylase secreted by the methylotrophic yeast pichia pastoris: comparison with wild-type recombinant glucoamylase produced using saccharomyces cerevisiae and aspergillus niger as hosts. | glucoamylase from aspergillus niger (identical to aspergillus awamori glucoamylase) is an industrially important, multidomain n- and o-glycosylated starch-hydrolase. to produce protein-engineered glucoamylase, heterologous expression is established in the methylotrophic yeast pichia pastoris. using the vector phil-d2, the cdna encoding a. awamori glucoamylase is inserted in the yeast genome downstream of the 5' aox1 promoter to replace the aox1 gene. induction by 0.75% methanol for 48 h led to s ... | 1997 | 9056481 |
| expression in pichia pastoris and purification of aspergillus awamori glucoamylase catalytic domain. | in this paper we report the expression in pichia pastoris, purification, and characterization of the aspergillus awamori glucoamylase catalytical domain (gac). pichia pastoris produced gac to the level of 0.4 g per liter medium. this production level is about the same level as that gained for recombinant ga from aspergillus and about 100-fold more than previously achieved by saccharomyces cerevisiae. the gac expressed in pichia pastoris was purified by two independent chromatographic methods emp ... | 1997 | 9179293 |
| cloning and characterization of two rhamnogalacturonan hydrolase genes from aspergillus niger. | a rhamnogalacturonan hydrolase gene of aspergillus aculeatus was used as a probe for the cloning of two rhamnogalacturonan hydrolase genes of aspergillus niger. the corresponding proteins, rhamnogalacturonan hydrolases a and b, are 78 and 72% identical, respectively, with the a. aculeatus enzyme. in a. niger cultures which were shifted from growth on sucrose to growth on apple pectin as a carbon source, the expression of the rhamnogalacturonan hydrolase a gene (rhga) was transiently induced afte ... | 1997 | 9212401 |
| expression and functional analysis of a hyperglycosylated glucoamylase in a parental host, aspergillus awamori var. kawachi. | a modified glucoamylase gene (glaa) with an extra thr- and ser-rich gp-i domain (t. semimaru, m. goto, k. furukawa, and s. hayashida, appl. environ. microbiol. 61:2885-2890, 1995) was introduced into a mutant parental host, aspergillus awamori var. kawachi, in which the original glaa gene had been completely deleted and replaced with the hygromycin phosphotransferase gene. the modified glaa was successfully expressed and secreted. the modified glucoamylase possessed higher digestibility of raw c ... | 1997 | 9212440 |
| an aspergillus awamori acetylesterase: purification of the enzyme, and cloning and sequencing of the gene. | an inducible acetylesterase was purified from the culture medium of aspergillus awamori strain ifo4033 growing on wheat-bran culture by ion-exchange, gel-filtration and hydrophobic-interaction chromatographies. the purified enzyme had an mr of 31000 and contained asn-linked oligosaccharides. the enzyme liberated acetic acid from wheat bran, hydrolysed only alpha-naphthyl acetate and propionate when aromatic esters were used for the substrate, and was tentatively classified as a carboxylic estera ... | 1997 | 9291122 |
| characterization of the bip gene of aspergillus awamori encoding a protein with an hdel retention signal homologous to the mammalian bip involved in polypeptide secretion. | a dna fragment containing an open reading frame of 2016 nucleotides has been cloned from the dna of aspergillus awamori by hybridization with a probe internal to the kar2 (bip) gene of saccharomyces cerevisiae. the 73.4-kda-encoded protein showed very high similarity to the endoplasmic reticulum (er) lumenal bip protein of s. cerevisiae, kluyveromyces lactis, schizosaccharomyces pombe, and animal and plant cells. the bip protein contains a polar n-terminal end followed by a 18-amino-acid strongl ... | 1997 | 9294262 |
| the er chaperone encoding bipa gene of black aspergilli is induced by heat shock and unfolded proteins. | we describe the cloning and characterisation of the bip gene homologues of the filamentous fungi aspergillus niger and aspergillus awamori. the bip genes of these black aspergilli encode an identical protein of 672 amino acids, which has a high homology with the bip protein from saccharomyces cerevisiae and contains a putative signal sequence of 38 amino acids. the dna sequences of the aspergillus bip genes diverge in particular in the three intronic sequences and the 5'- and 3'- noncoding regio ... | 1997 | 9370263 |
| minimizing nonproductive substrate binding: a new look at glucoamylase subsite affinities. | a subsite model as proposed by hiromi [hiromi, k. (1970) biochem. biophys. res. commun. 40, 1-6] has been applied to various hydrolases including glucoamylase (ga). the model assumes a single enzyme complex, a hydrolytic rate constant which is independent of substrate length, and a ratelimiting hydrolytic step. recent kinetic studies with ga contradict these assumptions. here we reevaluate the substrate binding of ga studying the pre-steady-state kinetics with glucose, which is reported here for ... | 1997 | 9398219 |
| detection of aspergillus ribosomal rna using biotinylated oligonucleotide probes. | aspergillosis continues to be a devastating disease entity that results in significant mortality in immunosuppressed patients. rapid diagnosis is often required to initiate appropriate therapy. although the histopathologist may be able to visualize fungal organisms in tissue specimens, the histology of aspergillus species may overlap with a variety of fungi, so diagnosis often relies on fungal cultures that can take weeks to complete. recently, an in situ hybridization assay targeting aspergillu ... | 1997 | 9458383 |
| influence of the process parameters on the morphology and enzyme production of aspergilli. | several papers have been published dealing with various fungi to determine their morphology, enzyme production or process performance. however, no publication considered all of these aspects simultaneously. in the case of the production of xylanase by aspergillus awamori the interrelationship of various key parameters are investigated. the influence of the reactor type (shake flasks, stirred tank and airlift tower loop reactor), the medium composition (semisynthetic and complex medium with wheat ... | 1998 | 9468803 |
| effect of introducing proline residues on the stability of aspergillus awamori. | in aspergillus awamori glucoamylase, ala27, ala393, ala435, ser436 and ser460 were replaced with proline residues, in order to stabilize the enzyme by forming more rigid peptide backbones. specific activities were unaffected except for a decrease in ser460-->pro glucoamylase. thermostability was increased in ser436-->pro glucoamylase, unchanged in ala435-->pro glucoamylase and decreased in ala27-->pro, ala393-->pro glucoamylases. as measured by circular dichroism, mutant glucoamylases ala435-->p ... | 1997 | 9488144 |
| restoration of catalytic activity beyond wild-type level in glucoamylase from aspergillus awamori by oxidation of the glu400-->cys catalytic-base mutant to cysteinesulfinic acid. | glucoamylase catalyzes the hydrolysis of glucosidic bonds with inversion of the anomeric configuration. site-directed mutagenesis and three-dimensional structure determination of the glucoamylase from aspergillus awamori previously identified glu179 and glu400 as the general acid and base catalyst, respectively. the average distance between the two carboxyl groups was measured to be 9.2 a, which is typical for inverting glycosyl hydrolases. in the present study, this distance was increased by re ... | 1998 | 9521693 |
| enzymatic properties of the cysteinesulfinic acid derivative of the catalytic-base mutant glu400-->cys of glucoamylase from aspergillus awamori. | the pka of the catalytic base was lowered and its distance to the general acid catalyst, glu179, was increased in the glucoamylase from aspergillus awamori by replacing the catalytic base glu400 with cysteine followed by oxidation to cysteinesulfinic acid [fierobe, h.-p., mirgorodskaya, e., mcguire, k. a., roepstorff, p., svensson, b. and clarke, a. j. (1998) biochemistry 37, 3743-3752. 1h nmr spectroscopy demonstrated that the oxidized mutant glu400-->cys-so2h glucoamylase, like the wild-type, ... | 1998 | 9521694 |
| influence of morphology on product formation in aspergillus awamori during submerged fermentations | the relationship between fungal morphology and heterologous protein production was examined for an aspergillus awamori strain during a series of fermentations with a batch phase followed by a fed-batch phase. agitation rate and inoculation concentration were used as controlled variables to generate different fungal morphologies in 20-dm3 stirred tank reactors. morphology was quantitatively characterized using image analysis. the different agitation rates and inoculum concentrations had large eff ... | 1998 | 9548774 |
| saccharification and alcohol fermentation in starch solution of steam-exploded potato. | steam explosion pretreatment of potato for the efficient production of alcohol was experimentally studied. the amount of water-soluble starch increased with the increase of steam pressure, but the amounts of methanol-soluble material and klason lignin remained insignificant, regardless of steam pressure. the potatoes exploded at high pressure were hydrolyzed into a low molecular liquid starch, and then easily converted into ethanol by simultaneous saccharification and fermentation using mixed mi ... | 1998 | 9554083 |
| mutations to alter aspergillus awamori glucoamylase selectivity. i. tyr48phe49-->trp, tyr116-->trp, tyr175-->phe, arg241-->lys, ser411-->ala and ser411-->gly. | glucoamylase mutations to reduce isomaltose formation from glucose condensation and thus increase glucose yield from starch hydrolysis were designed to produce minor changes in the active site at positions not totally conserved. tyr175-->phe and ser411-->gly glucoamylases had catalytic efficiencies on dp 2-7 maltooligosaccharides like those of wild-type glucoamylase, while the catalytic efficiencies of tyr116-->trp, arg241-->lys and ser411-->ala glucoamylases were reduced by about half and tyr48 ... | 1998 | 9605546 |
| mutations to alter aspergillus awamori glucoamylase selectivity. ii. mutation of residues 119 and 121. | mutations ser119-->glu, ser119-->gly, ser119-->trp, gly121-->ala and gly121-->ala/ser411-->gly were constructed in glucoamylase to change substrate specificity. mutation ser411-->gly was already known to decrease glucoamylase selectivity toward isomaltose formation and to increase peak glucose yield. all mutated glucoamylases had slightly lower specific activities on maltose than on wild-type glucoamylase. ser119-->glu, ser119-->gly and ser119-->trp glucoamylases were about as active on isomalto ... | 1998 | 9605547 |
| secretion of the sweet-tasting protein thaumatin by recombinant strains of aspergillus niger var. awamori. | a recombinant form of the sweet-tasting protein thaumatin has been produced in the filamentous fungus aspergillus niger var. awamori. expression cassettes containing a synthetic gene encoding thaumatin ii were prepared and used to transform aspergillus niger var. awamori strain nrrl312. several fungal strains capable of synthesizing and secreting thaumatin into the culture medium were generated, and their production capabilities were determined, first in shake flasks and later in a laboratory fe ... | 1998 | 9615480 |
| a system for production of commercial quantities of human lactoferrin: a broad spectrum natural antibiotic. | we previously reported the production of limited quantities of biologically active recombinant human lactoferrin in the filamentous fungus aspergillus oryzae. in the present study, we report a modification of this production system combined with a classical strain improvement program that has enabled production of levels of recombinant human lactoferrin in excess of 2 g/l. the protein was expressed in aspergillus awamori as a glucoamylase fusion polypeptide which was secreted into the growth med ... | 1995 | 9634791 |
| protein engineering of aspergillus awamori glucoamylase to increase its ph optimum. | to increase the ph optimum of glucoamylase (ga), five mutations-s411g, s411a, s411c, s411h and s411d--were designed to destabilize the carboxylate ion form of glu400, the catalytic base, by removing or weakening the hydrogen bond between ser411 and glu400, and thereby raising its pk. the substitution of alanine, histidine and aspartate were also designed to study the additional effects of polarity and both positive and negative charges, respectively. s411g ga had catalytic efficiencies like thos ... | 1998 | 9681871 |
| mutations to alter aspergillus awamori glucoamylase selectivity. iii. asn20-->cys/ala27-->cys, ala27-->pro, ser30-->pro, lys108-->arg, lys108-->met, gly137-->ala, 311-314 loop, tyr312-->trp and ser436-->pro. | mutations asn20-->cys/ala27-->cys (ss), ala27-->pro, ser30-->pro, lys108-->arg, gly137-->ala, tyr312-->trp and ser436-->pro in aspergillus awamori glucoamylase, along with a mutation inserting a seven-residue loop between tyr311 and gly314 (311-314 loop), were made to increase glucose yield from maltodextrin hydrolysis. no active lys108-->met glucoamylase was found in the supernatant after being expressed from yeast. lys108-->arg, 311-314 loop and tyr312-->trp glucoamylases have lower activities ... | 1998 | 9681872 |
| characterization and nitrogen-source regulation at the transcriptional level of the gdha gene of aspergillus awamori encoding an nadp-dependent glutamate dehydrogenase. | a 28.7-kb dna region containing the gdha gene of aspergillus awamori was cloned from a genomic dna library. a fragment of 2570 nucleotides was sequenced that contained orf1, of 1380 bp, encoding a protein of 460 amino acids (mr 49.4 kda). the encoded protein showed high similarity to the nadp-dependent glutamate dehydrogenases of different organisms. the cloned gene was functional since it complemented two different aspergillus nidulans gdha mutants, restoring high levels of nadp-dependent gluta ... | 1998 | 9683675 |
| expression and secretion of defined cutinase variants by aspergillus awamori. | several cutinase variants derived by molecular modelling and site-directed mutagenesis of a cutinase gene from fusarium solani pisi are poorly secreted by saccharomyces cerevisiae. the majority of these variants are successfully produced by the filamentous fungus aspergillus awamori. however, the l51s and t179y mutations caused reductions in the levels of extracellular production of two cutinase variants by a. awamori. metabolic labelling studies were performed to analyze the bottleneck in enzym ... | 1998 | 9687432 |
| structures of enzymically derived oligosaccharides from sorghum glucuronoarabinoxylan. | oligosaccharides derived from alkali-extracted sorghum glucuronoarabinoxylan by digestion with a combination of (1-->4)-beta-d-arabinoxylan arabinofuranohydrolase (axh) and endo-(1-->4)-beta-d-xylanase (xyl i), both from aspergillus awamori, were purified by size-exclusion chromatography followed by preparative high-performance anion-exchange chromatography. structural studies including monosaccharide analysis, methylation analysis, 1h nmr spectroscopy, and mass spectrometry were carried out, re ... | 1998 | 9691450 |
| agrobacterium tumefaciens-mediated transformation of filamentous fungi. | agrobacterium tumefaciens transfers part of its ti plasmid, the t-dna, to plant cells during tumorigenesis. it is routinely used for the genetic modification of a wide range of plant species. we report that a. tumefaciens can also transfer its t-dna efficiently to the filamentous fungus aspergillus awamori, demonstrating dna transfer between a prokaryote and a filamentous fungus. we transformed both protoplasts and conidia with frequencies that were improved up to 600-fold as compared with conve ... | 1998 | 9743116 |
| effect on thermostability and catalytic activity of introducing disulfide bonds into aspergillus awamori glucoamylase. | two additional disulfide bonds and three combined thermostabilizing mutations were introduced into aspergillus awamori glucoamylase to test their effects on enzyme thermostability and catalytic properties. the single cysteine mutations n20c, a27c, t72c and a471c were made and combined to produce the double cysteine mutations n20c/ a27c and t72c/a471c. the double cysteine mutants were expressed efficiently in saccharomyces cerevisiae, and disulfide bonds formed spontaneously after fermentation. a ... | 1998 | 9749918 |
| purification and substrate specificities of two alpha-l-arabinofuranosidases from aspergillus awamori ifo 4033. | alpha-l-arabinofuranosidases i and ii were purified from the culture filtrate of aspergillus awamori ifo 4033 and had molecular weights of 81,000 and 62,000 and pis of 3.3 and 3.6, respectively. both enzymes had an optimum ph of 4.0 and an optimum temperature of 60 degreesc and exhibited stability at ph values from 3 to 7 and at temperatures up to 60 degrees c. the enzymes released arabinose from p-nitrophenyl-alpha-l-arabinofuranoside, o-alpha-l-arabinofuranosyl-(1-->3)-o-beta-d-xylopyranosyl-( ... | 1998 | 9758835 |
| stabilization of aspergillus awamori glucoamylase by proline substitution and combining stabilizing mutations. | to stabilize aspergillus awamori glucoamylase (ga), three proline substitution mutations were constructed. when expressed in saccharomyces cerevisiae, ser30-->pro (s30p) stabilized the enzyme without decreased activity, whereas asp345-->pro (d345p) did not significantly alter and glu408-->pro (e408p) greatly decreased enzyme thermostability. the s30p mutation was combined with two previously identified stabilizing mutations: gly137-->ala, and asn20-->cys/ala27-->cys (which creates a disulfide bo ... | 1998 | 9796827 |
| effects of mutations in the starch-binding domain of bacillus macerans cyclodextrin glycosyltransferase. | cyclodextrin glycosyltransferase (cgtase) is an industrially important enzyme that produces cyclodextrins (cd) from starch by intramolecular transglycosylation. cgtase consists of five globular domains labeled a through e. to better understand the role of domain e in cgtase catalysis, we have constructed several mutants of bacillus macerans cgtase. removing the entire e domain resulted in an inactive enzyme. adding six amino acids between domains d and e caused a decrease in activity and thermos ... | 1998 | 9828462 |
| analysis of the role of the gene bipa, encoding the major endoplasmic reticulum chaperone protein in the secretion of homologous and heterologous proteins in black aspergilli. | the function of the endoplasmic-reticulum-localized chaperone binding protein (bip) in relation to protein secretion in filamentous fungi was studied. it was shown that the overproduction of several homologous and heterologous recombinant proteins by aspergillus strains induces the expression of bipa, the bip-encoding gene from aspergillus niger and aspergillus awamori. as this result could imply that bip plays a role in protein overproduction, the effect of modulation of bipa gene expression on ... | 1998 | 9830095 |
| purification and characterization of a feruloylesterase from aspergillus awamori. | a feruloylesterase was purified from the extracellular broth of aspergillus awamori grown on wheat bran culture. the purified enzyme gave a single band on sds-polyacrylamide gel electrophoresis and isoelectric focusing, with an apparent m(r) of 35,000 and a pi of 3.8, respectively. the substrate specificity of the purified enzyme differed obviously from that of acetylesterase of a. awamori. the enzyme bound to microcrystalline cellulose. | 1998 | 9836439 |
| thaumatin production in aspergillus awamori by use of expression cassettes with strong fungal promoters and high gene dosage. | four expression cassettes containing strong fungal promoters, a signal sequence for protein translocation, a kex protease cleavage site, and a synthetic gene (tha) encoding the sweet protein thaumatin ii were used to overexpress this protein in aspergillus awamori lpr66, a pepa protease-deficient strain. the best expression results were obtained with the gdha promoter of a. awamori or with the gpda promoter of aspergillus nidulans. there was good correlation of tha gene dosage, transcript levels ... | 1999 | 10049878 |
| structure of recombinant human lactoferrin expressed in aspergillus awamori. | human lactoferrin (hlf) has considerable potential as a therapeutic agent. overexpression of hlf in the fungus aspergillus awamori has resulted in the availability of very large quantities of this protein. here, the three-dimensional structure of the recombinant hlf has been determined by x-ray crystallography at a resolution of 2.2 a. the final model, comprising 5339 protein atoms (residues 1-691, 294 solvent molecules, two fe3+and two co32- ions), gives an r factor of 0.181 (free r = 0.274) af ... | 1999 | 10089347 |
| effects of dissolved oxygen tension and mechanical forces on fungal morphology in submerged fermentation. | the effects of dissolved oxygen tension and mechanical forces on fungal morphology were both studied in the submerged fermentation of aspergillus awamori. pellet size, the hairy length of pellets, and the free filamentous mycelial fraction in the total biomass were found to be a function of the mechanical force intensity and to be independent of the dissolved oxygen tension provided that the dissolved oxygen tension was neither too low (5%) nor too high (330%). when the dissolved oxygen concentr ... | 1998 | 10099217 |
| intrinsic kinetic parameters of the pellet forming fungus aspergillus awamori | fungi like aspergillus awamori may spontaneously form pellets, which introduces an extra oxygen transfer resistance and influences the activity of the microorganism. consequently, dramatic variations of apparent kinetics are reported in literature, due to variations in culture conditions, e.g., oxygen bulk concentration and pellet morphology. true intrinsic growth parameters like maximum growth rate and biomass yield, are important for process modelling and design. values for these parameters ma ... | 1998 | 10099283 |
| functional analysis of o-linked oligosaccharides in threonine/serine-rich region of aspergillus glucoamylase by expression in mannosyltransferase-disruptants of yeast. | the glaa gene encoding glucoamylase i (gai) of aspergillus awamori var. kawachi was heterologously expressed in mannosyltransferase mutants of saccharomyces cerevisiae, in which the pmt1 gene and the kre2 gene were disrupted. the gai enzymes expressed in these yeast mutant cells exhibited a lesser extent of o-glycosylation. secretion of gai expressed in the pmt1-disruptant and in the kre2-disruptant, respectively, was almost the same as that of gai expressed in wild type (wt) strains. the number ... | 1999 | 10102986 |
| mutations to alter aspergillus awamori glucoamylase selectivity. iv. combinations of asn20-->cys/ala27-->cys, ser30-->pro, gly137-->ala, 311-4 loop, ser411-->ala and ser436-->pro. | six previously constructed and nine newly constructed aspergillus awamori glucoamylases with multiple mutations made by combining existing single mutations were tested for their ability to produce glucose from maltodextrins. multiple mutations have cumulative effects on glucose yield, specific activity and thermostability. no general correlation between glucose yield and thermostability was observed, although mutations that presumably impede unfolding at high temperatures uniformly increase ther ... | 1999 | 10195288 |
| influence of microbial concentration on the rheology of non-newtonian fermentation broths. | the objective of this study was to quantify the effect of fungal biomass concentration on the rheology of non-newtonian fermentation systems. batch fermentations of penicillium chrysogenum were carried out with glucose as the sole carbon source. the flow behavior of the system was characterized at various fermentation times and was adequately described by the power-law model. the apparent viscosity of the fermentation broth was significantly affected by biomass concentrations in the fermenter. f ... | 1999 | 10222579 |
| transformation of aspergillus awamori by agrobacterium tumefaciens-mediated homologous recombination. | agrobacterium tumefaciens is known to transfer part of its tumor-inducing (ti) plasmid to the filamentous fungus aspergillus awamori by illegitimate recombination with the fungal genome. here, we show that when this ti dna shares homology with the a. awamori genome, integration can also occur by homologous recombination. on the basis of this finding, we have developed an efficient method for constructing recombinant mold strains free from bacterial dna by a. tumefaciens-mediated transformation. ... | 1999 | 10385327 |
| improving operating performance of glucoamylase by mutagenesis. | the potential operating temperature of aspergillus awamori glucoamylase has been increased by several thermostable mutations that reduce irreversible thermoinactivation. other mutations have been isolated that increase selectivity of alpha-1,4 over alpha-1,6 glycosidic bonds, resulting in fewer alpha-1,6 linked reversion products, thus increasing glucose yield. interestingly, many thermostable mutations also increase selectivity and yields, suggesting that enzyme flexibility plays a role in acco ... | 1999 | 10449316 |
| efficient utilization of starch by a recombinant strain of saccharomyces cerevisiae producing glucoamylase and isoamylase. | two plasmids, designated prti and pti, were constructed to allow the integration of a bacterial isoamylase gene (iso) into saccharomyces cerevisiae g23-8 chromosome. the integrative plasmid prti comprises the iso gene from pseudomonas amyloderamosa, a portion of s. cerevisiae ribosomal dna (rdna), s. cerevisiae trp1 gene deficient in promoter and the bacterial vector psp72. the structure of plasmid pti is similar to that of prti, except that it lacks an rdna segment. the aspergillus awamori gluc ... | 2000 | 10669402 |
| molecular characterization of a pdi-related gene prpa in aspergillus niger var. awamori. | a gene (prpa) homologous to the protein disulfide isomerase gene was isolated from aspergillus niger by southern hybridization using the pdi1 gene isolated from trichoderma reesei as a dna probe. the corresponding cdna of the prpa gene has also been isolated from an a. niger var. awamori cdna library. the prpa gene does not belong to any currently recognized family of protein disulfide isomerases since it contains only a single conserved thioredoxin domain at the n-terminus of the protein. the c ... | 2000 | 10672446 |
| physiological aspects of fungi isolated from root nodules of faba bean (vicia faba l.). | the present study was made to isolate and assess some physiological characteristics of root nodule-colonizing fungi. during this study, 17 fungal species were isolated from root nodule samples taken from faba bean plants (vicia faba l.) collected from different sites at assiut area (egypt). the growth of faba bean plants in pots was significantly promoted by soil inoculation with most fungi. growth was checked in pots with inocula of cladosporium cladosporioides, fusarium moniliforme, f: oxyspor ... | 2000 | 10772156 |
| soybean meal fermented by aspergillus awamori increases the cytochrome p-450 content of the liver microsomes of mice. | the effect of soybean meal fermented by aspergillus awamori on the acute lethality of acetaldehyde, pentobarbital sleeping time, and cytochrome p-450 content of the hepatic microsomes was studied in mice. most of the daidzin and genistin in soybean meal (sbm) were converted into the respective aglycones, daidzein and genistein, by fermentation. in experiment 1, mice were fed isonitrogenic test diets with one of the following five protein sources for 28 d: casein, sbm, fermented and hot-air-dried ... | 2000 | 10775399 |
| penicillopepsin-jt2, a recombinant enzyme from penicillium janthinellum and the contribution of a hydrogen bond in subsite s3 to k(cat). | the nucleotide sequence of the gene (pepa) of a zymogen of an aspartic proteinase from penicillium janthinellum with a 71% identity in the deduced amino acid sequence to penicillopepsin (which we propose to call penicillopepsin-jt1) has been determined. the gene consists of 60 codons for a putative leader sequence of 20 amino acid residues, a sequence of about 150 nucleotides that probably codes for an activation peptide and a sequence with two introns that codes for the active aspartic proteina ... | 2000 | 10850809 |
| bioaffinity extraction of glucoamylase in aqueous two-phase systems using starch as free bioligand. | aqueous two-phase systems with bioligands bound to the polymer, usually polyethylene glycol (peg), have high selectivity. however, such kind of systems can be costly since some specific synthetic ligands are expensive, making them non-viable for bioaffinity extraction of low-cost proteins. the use of free bioligands is an alternative for decreasing the high cost of these systems. this work describes the use of starch as a free bioligand on the partition of glucoamylase in peg 300-phosphate syste ... | 2000 | 10942292 |
| purification and characterization of acid-stable protopectinase produced by aspergillus awamori in solid-state fermentation. | aspergillus awamori ifo 4033 produced an acid-stable protopectinase in solid-state fermentation using wheat bran as the medium. the enzyme was purified to a homogeneous preparation with anion-exchange, hydrophobic, and size-exclusion chromatography. the enzyme was a monomeric protein of 52 kda, by sds-page analysis, with an isoelectric point of ph 3.7. the optimum ph for enzyme activity was 2.0, and it was most active at 50 degrees c (at ph 2.0) and was stable up to 50 degrees c (at ph 2.0). the ... | 2000 | 10945248 |
| cloning and heterologous expression of gene encoding a polygalacturonase from aspergillus awamori. | a polygalacturonase gene of aspergillus awamori ifo 4033 was cloned by genomic southern hybridization with a probe of a dna fragment synthesized by pcr. this was done using primers constructed based on the n-terminal amino acid sequence of a polygalacturonase, protopectinase-as, produced by the strain and the consensus internal amino acid sequence of fungal polygalacturonases. the cloned polygalacturonase gene, containing an orf, encodes 362 amino acids, including a 52-bp intron. it contains the ... | 2000 | 10993142 |
| purification and characterization of an endo-polygalacturonase. from aspergillus awamori. | an extracellular endo-polygalacturonase (pgase) produced by aspergillus awamori ifo 4033 was isolated from the culture filtrate. the enzyme was purified to a homogeneous preparation with cation-exchange and size-exclusion chromatographies. its properties were investigated, comparing them with that of recombinant pgx2 gene product, a pgase having protopectinase activity. this enzyme was a monomeric protein of 41 kda, with an isoelectric point of ph 6.1. the characteristics of this pgase substanti ... | 2000 | 10993164 |
| replacement and deletion mutations in the catalytic domain and belt region of aspergillus awamori glucoamylase to enhance thermostability. | three single-residue mutations, asp71-->asn, gln409-->pro and gly447-->ser, two long-to-short loop replacement mutations, gly23-ala24-asp25-gly26-ala27-trp28- val29-ser30-->asn-pro-pro (23-30 replacement) and asp297-ser298-glu299-ala300-val301-->ala-g ly-ala (297-301 replacement) and one deletion mutation removing glu439, thr440 and ser441 (delta439-441), all based on amino acid sequence alignments, were made to improve aspergillus awamori glucoamylase thermostability. the first and second singl ... | 2000 | 11054460 |
| glucoamylase: structure/function relationships, and protein engineering. | glucoamylases are inverting exo-acting starch hydrolases releasing beta-glucose from the non-reducing ends of starch and related substrates. the majority of glucoamylases are multidomain enzymes consisting of a catalytic domain connected to a starch-binding domain by an o-glycosylated linker region. three-dimensional structures have been determined of free and inhibitor complexed glucoamylases from aspergillus awamori var. x100, aspergillus niger, and saccharomycopsis fibuligera. the catalytic d ... | 2000 | 11150611 |
| overexpression and lack of degradation of thaumatin in an aspergillopepsin a-defective mutant of aspergillus awamori containing an insertion in the pepa gene. | a gene encoding the sweet-tasting protein thaumatin (tha) with optimized codon usage was expressed in aspergillus awamori. mutants of a. awamori with reduced proteolytic activity were isolated. one of these mutants, named lpr66, contained an insertion of about 200 bp in the pepa gene, resulting in an inactive aspergillopepsin a. in vitro thaumatin degradation tests confirmed that culture broths of mutant lpr66 showed only a small thaumatin-degrading activity. a. awamori lpr66 has been used as ho ... | 2000 | 11152068 |
| copper accumulation by aspergillus awamori. | aspergillus awamori accumulated cu2+ from aqueous solutions. the level of copper uptake was dependent on the ambient metal concentration. the process consisted of two phases: a fast initial phase and a slower secondary phase. chelation of these ions occurs by chemical, equilibrated and saturable mechanism, following the mathematical models of langmuir and freundlich, with better performance on the langmuir model. data transformation allowed us to calculate the kinetic constants of the sorption r ... | 2000 | 11271803 |
| [immobilization of glucoamylase from aspergillus awamori 466, and properties of the preparation]. | glucoamylase from aspergillus awamori 466 was immobilized on various supports. the enzyme sorption depends on its amount, the type of support, and immobilization conditions. the kinetics of acidic inactivation of the native and immobilized enzyme was studied. the immobilized enzyme was more resistant to temperature and ph. the mechanism of the enzyme binding to the support was investigated by ir spectroscopy. | 2001 | 11357426 |
| cloning of a phenol oxidase gene from acremonium murorum and its expression in aspergillus awamori. | fungal multicopper oxidases have many potential industrial applications, since they perform reactions under mild conditions. we isolated a phenol oxidase from the fungus acremonium murorum var. murorum that was capable of decolorizing plant chromophores (such as anthocyanins). this enzyme is of interest in laundry-cleaning products because of its broad specificity for chromophores. we expressed an a. murorum cdna library in saccharomyces cerevisiae and subsequently identified enzyme-producing ye ... | 2001 | 11375170 |
| production of beta-mannanase and beta-mannosidase from aspergillus awamori k4 and their properties. | beta-mannanase and beta-mannosidase from aspergillus awamori k4 was produced by solid culture with coffee waste and wheat bran. the optimum composition for enzyme production was 40% coffee waste-60% wheat bran. two enzymes were partially purified. optimum ph was about 5 for both enzymes, and optimum temperature was around 80 degrees c for beta-mannanase and 60-70 degrees c for beta-mannosidase. these enzymes produced some oligosaccharides from glucomannan and galactomannan by their hydrolyzing a ... | 2001 | 11381326 |
| identification, classification and phylogeny of the aspergillus section nigri inferred from mitochondrial cytochrome b gene. | the partial mitochondrial cytochrome b gene from 32 strains of 12 species belonging to aspergillus section nigri was amplified by the polymerase chain reaction and sequenced directly. using 402 nucleotide characters, nucleotide-based and amino acid-based phylogenetic trees were inferred and the genetic divergence among the species was evaluated. based on analyses of the 402-bp nucleotide and 133-amino acid sequences, strains were divided into 11 dna types and five amino acid types. aspergillus n ... | 2001 | 11425482 |
| factors affecting the production of a single-chain antibody fragment by aspergillus awamori in a stirred tank reactor. | a recombinant strain of aspergillus awamori expressing anti-lysozyme single chain antibody fragments (scfv), under the control of a xylanase promoter, was studied in order to investigate the impact of medium, induction regime and protease production on the expression of the product. experiments with the time of induction showed that the optimum results are achieved when induction is started in the late exponential phase (21 h after inoculation) improving the titer of the product from 14.5 mg l(- ... | 2001 | 11485420 |
| fungal flora of the digestive tract of 5 species of triatomines vectors of trypanosoma cruzi, chagas 1909. | a study of the mycobiota in the digestive tract of 5 important species of triatomines, triatoma brasiliensis, t infestans, t. sordida, t. pseudomaculata and t. vitticeps, was made. the digestive tracts of 164 adults and 535 nymphs of those triatomines were studied and 393 fungal strains were isolated. the genera with the greatest number of species were penicillium (19 species), aspergillus (17 species) and acremonium (5 species) and the most frequent species, in decreasing order, were penicilliu ... | 2001 | 11502063 |
| crystal structure of maltose phosphorylase from lactobacillus brevis: unexpected evolutionary relationship with glucoamylases. | maltose phosphorylase (mp) is a dimeric enzyme that catalyzes the conversion of maltose and inorganic phosphate into beta-d-glucose-1-phosphate and glucose without requiring any cofactors, such as pyridoxal phosphate. the enzyme is part of operons that are involved in maltose/malto-oligosaccharide metabolism. maltose phosphorylases have been classified in family 65 of the glycoside hydrolases. no structure is available for any member of this family. | 2001 | 11587643 |
| ethanol production from starch by immobilized aspergillus awamori and saccharomyces pastorianus using cellulose carriers. | a simultaneous saccharification and fermentation (ssf) process was investigated to produce ethanol using two kinds of cellulose carriers that were respectively suitable for immobilization of aspergillus awamori and saccharomyces pastorianus. the maximum ethanol concentration attained by the batch operation was 25.5 g l(-1). under suitable conditions, both cellulose carriers with immobilized cells could be reused efficiently for three cycles. the total amount of ethanol production was 66.0 g (per ... | 2001 | 11598811 |
| a defined level of protein disulfide isomerase expression is required for optimal secretion of thaumatin by aspegillus awamori. | thaumatin, a 22-kda protein containing eight disulfide bonds, is secreted by the filamentous fungus aspergillus awamori at levels which are dependent upon the extent of overexpression of protein disulfide isomerase (pdia). additional copies of the pdia-encoding gene pdia were introduced into a strain of a. awamori that expresses a cassette encoding thaumatin. transformants with different levels of pdia mrna and measured pdia levels were chosen for examination of the impact that pdia levels had o ... | 2001 | 11683266 |
| purification, characterization, gene cloning and preliminary x-ray data of the exo-inulinase from aspergillus awamori. | extracellular exo-inulinase has been isolated from a solid-phase culture of the filamentous fungus aspergillus awamori var. 2250. the apparent molecular mass of the monomer enzyme was 69 +/- kda, with a pi of 4.4 and a ph optimum of 4.5. the enzyme hydrolysed the beta-(2-->1)-fructan (inulin) and beta-(2-->6)-fructan (levan) via exo-cleavage, releasing fructose. the values for the michaelis constants k(m) and v(max) in the hydrolysis of inulin were 0.003 +/- 0.0001 mm and 175 +/- 5 micromol.min( ... | 2002 | 11829749 |
| polygalacturonase production by aspergillus awamori on wheat in solid-state fermentation. | the production of exo-polygalacturonase (exo-pg) and endo-pg by aspergillus awamori grown on wheat in solid-state fermentation was studied. endo- and exo-pg activities were detected after 24 h of inoculation. glucose released from starch hydrolysis acted as a catabolite repressor for the exo-pg enzyme. in contrast, endo-pg production was not affected by glucose repression. when milled grains were used, the particle-size distribution and the chemical composition of the medium influenced the rate ... | 2002 | 11876407 |
| effect of lactoferrin on helicobacter felis induced gastritis. | lactoferrin possesses antibiotic, antiinflammatory, and immune-modulating properties that may be active against the gastritis-, ulcer- and cancer-inducing bacterium helicobacter pylori. in vitro testing of bovine and human lactoferrin by several laboratories has shown significant bacteriostatic and bactericidal activity. subsequent in vivo testing of bovine lactoferrin in animal models of h. pylori infection has shown beneficial effects of this agent. our laboratory has utilized a mouse model th ... | 2002 | 11908634 |
| aminoadipate reductase gene: a new fungal-specific gene for comparative evolutionary analyses. | in fungi, aminoadipate reductase converts 2-aminoadipate to 2-aminoadipate 6-semialdehyde. however, other organisms have no homologue to the aminoadipate reductase gene and this pathway appears to be restricted to fungi. in this study, we designed degenerate primers for polymerase chain reaction (pcr) amplification of a large fragment of the aminoadipate reductase gene for divergent fungi. | 2002 | 11931673 |
| constitutive expression of the trichoderma reesei beta-1,4-xylanase gene (xyn2) and the beta-1,4-endoglucanase gene (egl) in aspergillus niger in molasses and defined glucose media. | the xylanase ii (xyn2)- and endoglucanase i (egl)-encoding regions of trichoderma reesei qm6a were successfully expressed in aspergillus niger d15 under the transcriptional control of the glyceraldehyde-6-phosphate dehydrogenase (gpd) promoter from a. niger and the glaa terminator of aspergillus awamori. a stable xyn2 transformant produced beta-xylanase activity of 8,000 nkat/ml and 5,000 nkat/ml in shake-flask cultures containing defined or 20% (v/v) molasses medium, respectively. the recombina ... | 2002 | 11954792 |
| xylanase production by aspergillus awamori in solid-state fermentation and influence of different nitrogen sources. | the use of purified xylan as a substrate for bioconversion into xylanases increases the cost of enzyme production. consequently, there have been attempts to develop a bioprocess to produce such enzymes using different lignocellulosic residues. filamentous fungi have been widely used to produce hydrolytic enzymes for industrial applications, including xylanases, whose levels in fungi are generally much higher than those in yeast and bacteria. considering the industrial importance of xylanases, th ... | 2001 | 11963896 |
| expression of an alpha-galactosidase from saccharomyces cerevisiae in aspergillus awamori and aspergillus oryzae. | a gene encoding alpha-galactosidase activity was isolated by polymerase chain reaction (pcr) from saccharomyces cerevisiae ncyc686 and separately placed under the control of transcriptional elements regulating alpha-amylase expression in aspergillus oryzae and glucoamylase expression in a. awamori. following transformation of both a. oryzae and a. awamori with their respective expression vectors, induction of heterologous alpha-galactosidase from positively selected clones was effected through t ... | 2002 | 12074058 |
| silencing of the aspergillopepsin b (pepb) gene of aspergillus awamori by antisense rna expression or protease removal by gene disruption results in a large increase in thaumatin production. | aspergillopepsin b was identified in culture broths of aspergillus awamori by in situ detection of its proteolytic activity and by immunodetection with anti-aspergillopepsin b antibodies. severe thaumatin degradation was observed after in vitro treatment of thaumatin with purified aspergillopepsin b. the pepb gene encoding aspergillopepsin b of a. awamori was cloned and characterized. it is located in chromosome iv of a. awamori, as shown by pulsed-field gel electrophoresis, and encodes a protei ... | 2002 | 12089041 |
| alcohol production from starch by mixed cultures of aspergillus awamori and immobilized saccharomyces cerevisiae at different agitation speeds. | the influence of different agitation speeds on alcoholic fermentation by free and immobilized cells of saccharomyces cerevisiae in a co-culture with free cells of aspergillus awamori was investigated. starch hydrolysis and glucose, glucoamylase and alpha-amylase accumulation in the fermentation medium was affected by agitation speed. maximum amounts of the enzymes were obtained at 150-200 rpm after 72 h where, maximum growth and glucose accumulations were noticed at 200-300 rpm. alcohol producti ... | 2002 | 12111743 |
| ultra scaledown to predict filtering centrifugation of secreted antibody fragments from fungal broth. | extracellularly expressed anti-hen egg lysozyme single-chain antibody fragments (scfv) produced by aspergillus awamori were recovered using filtering centrifugation. two filtering centrifuges with 0.5- and 30-l capacities were used to represent laboratory- and pilot-scale equipment, respectively. critical regime analysis using the computational fluid dynamics (cfd) technique provided information about the local energy dissipation rates in both units. experimental data indicated loss of scfv acti ... | 2002 | 12115401 |
| swollenin, a trichoderma reesei protein with sequence similarity to the plant expansins, exhibits disruption activity on cellulosic materials. | plant cell wall proteins called expansins are thought to disrupt hydrogen bonding between cell wall polysaccharides without hydrolyzing them. we describe here a novel gene with sequence similarity to plant expansins, isolated from the cellulolytic fungus trichoderma reesei. the protein named swollenin has an n-terminal fungal type cellulose binding domain connected by a linker region to the expansin-like domain. the protein also contains regions similar to mammalian fibronectin type iii repeats, ... | 2002 | 12199698 |
| effects of root-dip treatment with certain phosphate solubilizing microorganisms on the fusarial wilt of tomato. | root-dip application of bacillus subtilis, pseudomonas fluorescens, aspergillus awamori, aspergillus niger and penicillium digitatum resulted in significant decline in the rhizosphere population of fusarium oxysporum f. sp. lycopersici. a significant decrease in the severity of wilt occurred with a. awamori (37.1%) and p. digitatum (21.3%) compared to the control. root-dip treatment with the phosphate solubilizing microorganisms tested resulted in significant increase in the yield of tomato, bei ... | 2002 | 12227549 |
| evaluation of filamentous fungi and inducers for the production of endo-polygalacturonase by solid state fermentation. | aspergillus oryzae cct 3940, aspergillus awamori nrrl 3112 and a trichoderma sp.) were compared for their capacity to produce endo-polygalacturonase (endo-pg) in solid state fermentation. maximum pectinolytic activity was reached in 72 h of growth, the best two fungal strains being a. niger t0005007-2 and a. oryzae cct 3940. three types of commercial purified pectin and four of unprocessed pectin (tangerine, orange, tahiti lime and sweet lime rind) were used to assess the effect of pectin on the ... | 2002 | 12240994 |
| aspergillin pz, a novel isoindole-alkaloid from aspergillus awamori. | aspergillin pz was obtained from the fermentation of aspergillus awamori (nakazawa) by activity-guided fractionation and purification. its structure was elucidated on the basis of spectral data, especially by 2d nmr, and finally confirmed by an x-ray analysis. it could induce conidia of p. oryzae to deform moderately. | 2002 | 12374381 |