PMID(sorted descending)
pisatin resistance in dictyostelium discoideum and neurospora crassa: comparison of mutant phenotypes.the pea phytoalexin pisatin, at its inhibitory concentration, was shown to have two distinct inhibitory effects on amoebae of the cellular slime mould dictyostelium discoideum. one effect was cytolytic and was demonstrable even in non-growing cells whereas the second effect was observed only under conditions favourable to growth. pretreatment with a sublethal concentration of pisatin induced the amoebae to acquire resistance to both these effects. mutations in nysc that alter membrane sterols an ...19938126430
disruption of the gene coding for the 21.3-kda subunit of the peripheral arm of complex i from neurospora crassa.a 21.3-kda subunit of the peripheral arm of complex i from neurospora is encoded by a single chromosomal gene, nuo-21.3b. it is located on linkage group v of the fungal genome, linked to inl. we have isolated and characterized a genomic clone containing this nuclear gene. a dna fragment containing a portion of the coding region of the gene and upstream flanking sequences was introduced by transformation into a wild-type strain of neurospora crassa. a single copy transformant was selected and cro ...19948126004
disruption of the gene encoding the nadh-binding subunit of nadh: ubiquinone oxidoreductase in neurospora crassa. formation of a partially assembled enzyme without fmn and the iron-sulphur cluster this study, the gene of the 51-kda nadh-binding subunit of the mitochondrial nadh:ubiquinone oxidoreductase (complex i) in neurospora crassa was inactivated by homologous replacement with a defective gene copy. the resulting mutant, nuo51, lacks the 51-kda subunit and shows no complex i activity but still grows at one third of the wild-type growth rate. the enzyme activity of the alternative nadh:ubiquinone oxidoreductase(s) is increased twofold while the activities of the other mitochondrial ...19948125114
an efficient method for gene disruption in neurospora crassa.the frequency with which transforming dna undergoes homologous recombination at a chromosomal site can be quite low in some fungal systems. in such cases, strategies for gene disruption or gene replacement must either select against ectopic integration events or provide easy screening to identify homologous site, double-crossover insertion events. a protocol is presented for efficient isolation of neurospora crassa strains carrying a definitive null allele in a target gene. the protocol relies o ...19948121407
sequential gel mobility shift scanning of 5' upstream sequences of the neurospora crassa am (gdh) gene.we have used gel mobility shift assays to scan 1.7 kb of 5' non-coding sequence of the am (glutamate dehydrogenase) gene of neurospora crassa for binding by partially fractionated neurospora proteins. using genetic analysis this region had been shown to play an important role in the control of glutamate dehydrogenase (gdh) expression. gel mobility shift analysis identified three regions to which neurospora proteins bind specifically. two of these corresponded to the two elements previously defin ...19948121395
a crucial role of the mitochondrial protein import receptor mom19 for the biogenesis of mitochondria.the novel genetic method of "sheltered rip" (repeat induced point mutation) was used to generate a neurospora crassa mutant in which mom19, a component of the protein import machinery of the mitochondrial outer membrane, can be depleted. deficiency in mom19 resulted in a severe growth defect, but the cells remained viable. the number of mitochondrial profiles was not grossly changed, but mutant mitochondria were highly deficient in cristae membranes, cytochromes, and protein synthesis activity. ...19948120088
cytochalasin b-sensitive actin-mediated nuclear rna export in germinating conidia of neurospora crassa. 19938111347
a novel carotenoid biosynthesis gene coding for zeta-carotene desaturase: functional expression, sequence and phylogenetic origin.a dna fragment which has been isolated previously from an anabaena dna expression library was subcloned. the corresponding protein was overexpressed in escherichia coli. the recombinant enzyme was fully active in converting zeta-carotene into lycopene in vitro with neurosporene as an intermediate. a smaller fragment which still contained the active enzyme was sequenced. an open reading frame of 1497 bp was found coding for a protein consisting of 499 amino acids with the calculated molecular wei ...19948111038
small lipid-soluble cations are not membrane voltage probes for neurospora or saccharomyces.small lipid-soluble cations, such as tetraphenylphosphonium (tpp+) and tetraphenylarsonium (tpa+) are frequently used as probes of membrane voltage (delta psi, or vm) for small animal cells, organelles, and vesicles. because much controversy has accompanied corresponding measurements on 'walled' eukaryotic cells (plants, fungi), we studied their transport and relation to vm in the large-celled fungus neurospora crassa-where vm can readily be determined with microelectrodes-as well as in the most ...19948110820
cis,cis-muconate lactonizing enzyme from trichosporon cutaneum: evidence for a novel class of cycloisomerases in eucaryotes.the absolute stereochemical courses of cis,cis-muconate lactonizing enzyme (mle;ec from trichosporon cutaneum (tcmle) and chloromuconate cycloisomerase (mle ii; ec from pseudomonas sp b13 have been determined from 1h nmr measurements. both cycloisomerases convert cis,cis-muconate to (4s)-muconolactone by a syn lactonization, the absolute stereochemical outcome of which is identical to that observed with mle from pseudomonas putida. the regiochemical courses of cyclization of 3- ...19948110801
characterization of the mitochondrial processing peptidase of neurospora crassa.the mitochondrial processing peptidase (mpp) of neurospora crassa is constituted by an alpha- and a beta-subunit. we have purified alpha-mpp after expression in escherichia coli while beta-mpp was purified from mitochondria. a fusion protein between precytochrome b2 and mouse dihydrofolate reductase was expressed in e. coli, and the purified protein was used as substrate for mpp. both subunits of mpp are required for processing. mpp removes the matrix targeting signal of cytochrome b2 by a singl ...19948106471
physical mapping of meiotic crossover events in a 200-kb region of neurospora crassa linkage group i.we propose a general restriction fragment length polymorphism-based strategy to analyze the distribution of meiotic crossover events throughout specific genetic intervals. we have isolated 64 recombinant chromosomes carrying independent meiotic crossover events in the genetic interval eth-1-un-2 on linkage group i of neurospora crassa. thirty-eight crossover events were physically mapped with reference to a 200-kb region cloned by chromosome walking, using n. crassa lambda and cosmid libraries. ...19938104159
isolation and characterization of mutants defective in production of laccase in neurospora crassa.a protein synthesis inhibitor, cycloheximide, induces excretion of laccase in neurospora crassa. the lah-1 mutation results in excretion of a large amount of laccase even in the absence of cycloheximide. ten mutations were induced that suppress derepressed excretion of laccase in the lah-1 mutant. of these, seven second-site mutations were found to confer a laccase-noninducible phenotype, and were classified into two different complementation groups. four mutations define a locus designated lni- ...19938102779
the 12.3 kda subunit of complex i (respiratory-chain nadh dehydrogenase) from neurospora crassa: cdna cloning and chromosomal mapping of the gene.the 12.3 kda subunit of complex i (respiratory-chain nadh dehydrogenase) is a nuclear-coded protein of the hydrophobic fragment of the enzyme. we have isolated and sequenced a full-length cdna clone coding for this polypeptide. the deduced protein is 104 amino acid residues long with a molecular mass of 12305 da. this particular subunit of complex i lacks a cleavable mitochondrial targeting sequence. in agreement with its localization within complex i, we have found that this subunit behaves lik ...19938098209
the neurospora uvs-2 gene encodes a protein which has homology to yeast rad18, with unique zinc finger motifs.a clone containing the dna repair gene uvs-2 of neurospora crassa was identified from a neurospora genomic dna library using the sib-selection method. transformants were screened for resistance to methyl methane sulfonate (mms). a dna fragment that complements the uvs-2 mutation was subcloned by monitoring its ability to transform the uvs-2 mutant to mms resistance. deletion analysis of the cloned dna indicated that the size of the uvs-2 gene is approximately 1.6 kb. the identity of the uvs-2 ge ...19938097557
molecular cloning, characterization, and nucleotide sequence of nit-6, the structural gene for nitrite reductase in neurospora crassa.the neurospora crassa assimilatory nitrite reductase structural gene, nit-6, has been isolated. a cdna library was constructed from poly(a)+ rna isolated from neurospora mycelia in which nitrate assimilation had been induced. this cdna was ligated into lambda zap ii (stratagene) and amplified. this library was then screened with a polyclonal antibody specific for nitrite reductase. a total of six positive clones were identified. three of the six clones were found to be identical via restriction ...19938096840
pcr-based cloning of the full-length neurospora eukaryotic initiation factor 5a cdna: polyhistidine-tagging and overexpression for protein affinity binding.eukaryotic initiation factor 5a (eif-5a) is the only cellular protein known to contain a hypusine residue that is formed by transferring the aminobutyl moiety from spermidine to a specific lysine residue, followed by hydroxylation at the aminobutyl group. a simple pcr-based strategy was developed to obtain a full-length cdna of neurospora crassa eif-5a. the strategy consists of (i) the design of a pair of key primers (21-mer) based on the highly conserved eif-5a cdna domains known in other speci ...19948093005
the isolation and characterization of a neurospora crassa gene (ubi::crp-6) encoding a ubiquitin-40s ribosomal protein fusion protein.we have isolated and sequenced a neurospora crassa gene encoding a single copy of ubiquitin (ubi) fused to the s27a ribosomal (r) protein. we have opted to name this gene the ubiquitin/cytoplasmic r-protein gene 6 (ubi::crp-6). the ubi::crp-6 gene generates a 700-nucleotide (nt) transcript. it shares a 700-bp regulatory region with the cytoplasmic r-protein gene 5 (crp-5), a gene encoding the n. crassa s26 r-protein. the two genes are transcribed divergently from the common regulatory region and ...19948088539
suppressor mutants of neurospora crassa that tolerate allelic differences at single or at multiple heterokaryon incompatibility loci.allelic differences at any one of at least 11 heterokaryon incompatibility (het) loci in neurospora crassa trigger an incompatibility response: localized cell death at sites of hyphal anastomosis. we have isolated spontaneous and insertional suppressor mutants that are heterokaryon-compatible in spite of allelic differences at one or at several het loci. some intra- and extragenic mutants tolerated allelic differences only at single het loci. multi-tolerant spontaneous mutants were isolated by s ...19948088519
superoxide dismutase (sod-1) null mutants of neurospora crassa: oxidative stress sensitivity, spontaneous mutation rate and response to mutagens.enzymatic superoxide-dismutase activity is believed to be important in defense against the toxic effects of superoxide. although superoxide dismutases are among the best studied proteins, numerous questions remain concerning the specific biological roles of the various superoxide-dismutase types. in part, this is because the proposed damaging effects of superoxide are manifold, ranging from inactivation of certain metabolic enzymes to dna damage. studies with superoxide-deficient mutants have pr ...19948088518
functional analyses of the neurospora crassa mt a-1 mating type polypeptide.the neurospora crassa mt a-1 gene, encoding the mt a-1 polypeptide, determines a mating type properties: sexual compatibility and vegetative incompatibility with a mating type. we characterized in vivo and in vitro functions of the mt a-1 polypeptide and specific mutant derivatives. mt a-1 polypeptide produced in escherichia coli bound to specific dna sequences whose core was 5'-ctttg-3'. dna binding was a function of the mt a-1 hmg box domain (a dna binding motif found in high mobility group pr ...19948088517
isolation, sequence, and characterization of the cercospora nicotianae phytoene dehydrogenase gene.we have cloned and sequenced the cercospora nicotianae gene for the carotenoid biosynthetic enzyme phytoene dehydrogenase. analysis of the derived amino acid sequence revealed it has greater than 50% identity with its counterpart in neurospora crassa and approximately 30% identity with prokaryotic phytoene dehydrogenases and is related, but more distantly, to phytoene dehydrogenases from plants and cyanobacteria. our analysis confirms that phytoene dehydrogenase proteins fall into two groups: th ...19948085820
amino-acid alterations in the beta-tubulin gene of neurospora crassa that confer resistance to carbendazim and diethofencarb.we have previously shown that increased sensitivity to diethofencarb in the carbendazim(mbc)-resistant f914 strain of neurospora crassa is caused by a single amino-acid change in beta-tubulin, 198glu to gly. three diethofencarb-resistant mutants that are also resistant to mbc were isolated from strain f914. they contained single base-pair-substitution mutations in the beta-tubulin gene. the amino acid changes in beta-tubulin, phe from 250leu, val from 165ala, and ala from 237thr, were responsibl ...19948082187
gene inactivation in arabidopsis thaliana is not accompanied by an accumulation of repeat-induced point mutations.chromosomal integration of multicopy transgene inserts in higher plants is often followed by loss of expression. we have analysed whether this inactivation can trigger repeat-induced point mutations (rip) as has been observed in neurospora crassa. we have previously characterized transgenic lines of arabidopsis thaliana containing the hygromycin phosphotransferase (hpt) gene either as a unique sequence in plants expressing the gene, or as multimeric, closely linked repeats in clones that were re ...19948058043
cloning and characterization of the pepe gene of aspergillus niger encoding a new aspartic protease and regulation of pepe and pepc.we have cloned the pepe gene of aspergillus niger, encoding an aspartic protease (pepe), by screening a lambda genomic dna library with a heterologous probe, the neurospora crassa gene coding for a vacuolar proteinase. sequencing of pepe genomic and cdna clones revealed that the gene contains three introns, which are 91, 56 and 58-bp long. the deduced protein consists of 398 amino acids, has a putative signal sequence to allow transport into the endoplasmic reticulum and probably undergoes a sec ...19948056328
cell-surface receptors for gibbon ape leukemia virus and amphotropic murine retrovirus are inducible sodium-dependent phosphate symporters.cell surface receptors for gibbon ape leukemia virus (glvr-1) and murine amphotropic retrovirus (ram-1) are distinct but related proteins having multiple membrane-spanning regions. distant homology with a putative phosphate permease of neurospora crassa suggested that these receptors might serve transport functions. by expression in xenopus laevis oocytes and in mammalian cells, we have identified glvr-1 and ram-1 as sodium-dependent phosphate symporters. two-electrode voltage-clamp analysis ind ...19948041748
efficient independent activity of a monomeric, monofunctional dehydroquinate synthase derived from the n-terminus of the pentafunctional arom protein of aspergillus nidulans.the dehydroquinate synthase (dhq synthase) functional domain from the pentafunctional arom protein of aspergillus nidulans has previously been overproduced in escherichia coli [van den hombergh, moore, charles and hawkins (1992) biochem j. 284, 861-867]. we now report the purification of this domain to homogeneity and subsequent characterization. the monofunctional dhq synthase was found to retain efficient catalytic activity when compared with the intact pentafunctional arom protein of neurospo ...19948037684
cloning and characterization of the pho-2+ gene encoding a repressible alkaline phosphatase in neurospora crassa.the neurospora crassa phosphate-repressible alkaline phosphatase-encoding gene pho-2+ was cloned and its nucleotide sequence was determined. an open reading frame was found that contains four introns and encodes a putative protein of 555 amino acids. 'activator-independent expression' of ectopically integrated pho-2+ was observed, as noted before for ectopically integrated pho-4+.19948026754
atp synthase of yeast mitochondria. isolation of the f1 delta subunit, sequence and disruption of the structural gene.the delta-subunit was isolated from the purified yeast f1. partial protein sequences were determined by direct methods. from this information, degenerated primers were constructed. a part of the atp delta gene was amplified by polymerase chain reaction from yeast genomic dna. from the amplified dna sequence, a nondegenerated oligonucleotide probe was constructed to isolate a 2.6-kbp bamhi-ecori dna fragment bearing the whole gene. a 1036-bp drai fragment was sequenced. a 480-bp open reading fram ...19948026496
high yield expression of the neurospora crassa plasma membrane h(+)-atpase in saccharomyces cerevisiae.a simple system for high yield expression of the neurospora plasma membrane h(+)-atpase is described. two neurospora h(+)-atpase cdnas differing only in a few bases preceding the coding region were cloned into a high copy number yeast expression vector under the control of the constitutive promoter of the yeast plasma membrane h(+)-atpase, and the resulting plasmids were used to transform saccharomyces cerevisiae strain rs-72, which requires a plasmid-borne functional plasma membrane h(+)-atpase ...19948021283
the characterization of a mitochondrial acyl carrier protein isoform isolated from arabidopsis thaliana.a cdna clone was isolated from an arabidopsis leaf cdna library that shared a high degree of protein sequence identity with mitochondrial acyl carrier proteins (mtacps) isolated from neurospora crassa and bovine heart muscle. the cdna encoded an 88-amino acid mature protein that was preceded by a putative 35-amino acid presequence. in vitro protein import studies have confirmed that the presequence specifically targets this protein into pea mitochondria but not into chloroplasts. these studies i ...19948016262
mutants of neurospora crassa that alter gene expression and conidia development.several genes have been identified that are highly expressed during conidiation. inactivation of these genes has no observable phenotypic effect. transcripts of two such genes, con-6 and con-10, are normally absent from vegetative mycelia. to identify regulatory genes that affect con-6 and/or con-10 expression, strains were prepared in which the regulatory regions for these genes were fused to a gene conferring hygromycin resistance. mutants were then selected that were resistant to the drug dur ...19948016143
characterization of a membrane fragment of respiratory chain complex i from neurospora crassa. insights on the topology of the ubiquinone-binding site.1. a membrane fragment of complex i from the fungus neurospora crassa was isolated by immunoprecipitation from alkaline-extracted mitochondrial membranes. 2. analysis of the polypeptide composition of this hydrophobic domain of complex i has brought insights on the topology of two subunits of the enzyme, namely the 20.8 and 9.3 kda components. 3. our results indicate that the ubiquinone-binding site of complex i resides in the interface of the peripheral and membrane arms of the enzymes. the sig ...19948013735
interaction of phlorizin, a potent inhibitor of the na+/d-glucose cotransporter, with the nadph-binding site of mammalian catalases.phlorizin is a reversible inhibitor of the renal and small intestinal na+/d-glucose cotransporter. in an attempt to purify the na+/d-glucose cotransporter from a pig kidney brush border membrane fraction, we used an affi-gel affinity chromatography column to which 3-aminophlorizin had been coupled. a protein, composed according to crosslinking experiments of at least 3 subunits of molecular weight 60 kda, was found to bind specifically to the phlorizin column. this protein was subsequently ident ...19948003987
directed replacement of mt a by mt a-1 effects a mating type switch in neurospora test the functions of a mating type genes, we developed an efficient strategy to select transformants of neurospora crassa in which resident a mating type dna was replaced by cloned dna from the mt a idiomorph. cloned a idiomorphic dna could specify all functions, including fertility, of a mating type, but only when it replaced a dna at the mating type locus. only the mt a-1 region of the a idiomorph was necessary in order to specify a mating type. gene replacement events involved the homolog ...19948001795
spontaneous mutation at the mtr locus in neurospora: the molecular spectrum in wild-type and a mutator strain.sequence analysis of 34 mtr mutations has yielded the first molecular spectrum of spontaneous mutants in neurospora crassa. the great majority of the mutations are base substitutions (48%) or deletions (35%). in addition, sequence analysis of the entire mtr region, including the 1472-base pair open reading frame and 1205 base pairs of flanking dna, was performed in both the oak ridge and mauriceville strains of neurospora, which are known to be divergent at the dna level. sixteen sequence differ ...19948001794
identification and partial purification of calmodulin-binding microtubule-associated proteins from neurospora crassa.we have purified microtubule-associated proteins from neurospora crassa on the basis of heat stability and affinity to calmodulin. two proteins of molecular masses 170 kda and 190 kda have been partially purified. a third protein of 145 kda was purified almost to homogeneity, and we present evidence that this protein is a specific substrate for a ca2+/calmodulin-dependent protein kinase. the purified 170-, 190-, and 145-kda proteins induce the assembly of microtubules from purified porcine brain ...19948001548
fatty acid biosynthesis in novel ufa mutants of neurospora mutants of neurospora crassa having the ufa phenotype have been isolated. two of these mutants, like previously identified ufa mutants, require an unsaturated fatty acid for growth and are almost completely blocked in the de novo synthesis of unsaturated fatty acids. the new mutations map to a different chromosomal location than previously characterized ufa mutations. this implies that at least one additional genetic locus controls the synthesis of unsaturated fatty acids in neurospora.19948000539
cytochalasin b-sensitive actin-mediated nuclear rna export in germinating conidia of neurospora crassa. 19948000364
effect of carbon source and extracellular ph on the acidification of the culture medium and phosphatase excretion in neurospora crassa.exogenous ca2+ at concentrations up to 3.5 mm increases the sucrose-induced acidification of the culture medium when the mold neurospora crassa is grown on low-phosphate (pi) medium at ph 7.8. induction depends on the ph of the culture medium adjusted for conidial inoculation and on the absence of carbon sources generating cytoplasmic acetyl coa. furthermore, the excretion of pi-repressible acid and alkaline phosphatases was not stimulated by increasing exogenous ca2+ levels. we also provide evi ...19948000333
pulsed growth of fungal hyphal tips.somatic fungal hyphae are generally assumed to elongate at steady linear rates when grown under constant environmental conditions with ample nutrients. however, patterns of pulsed hyphal elongation were detected during apparent steady growth of hyphal tips in fungi from several major taxonomic groups (oomycetes, pythium aphanidermatum and saprolegnia ferax; zygomycetes, gilbertella persicaria; deuteromycetes, trichoderma viride; ascomycetes, neurospora crassa and fusarium culmorum; basidiomycete ...19947991610
polyamine metabolism and growth of neurospora strains lacking cis-acting control sites in the ornithine decarboxylase gene.ornithine decarboxylase (odc) initiates the synthesis of polyamines (putrescine, spermidine, and spermine) and is highly regulated. we wished to know the importance of the control of odc synthesis to the rates of growth and polyamine synthesis in the fungus, neurospora crassa. we identified two control sites of the spe-1 gene, encoding odc. one was an upstream activation region (uar) and the other was the dna encoding the long odc mrna leader, which governs polyamine-mediated repression of enzym ...19947979392
analysis of the dna-binding and dimerization activities of neurospora crassa transcription factor nuc-1.nuc-1, a positive regulatory protein of neurospora crassa, controls the expression of several unlinked target genes involved in phosphorus acquisition. the carboxy-terminal end of the nuc-1 protein has sequence similarity to the helix-loop-helix family of transcription factors. bacterially expressed and in vitro-synthesized proteins, which consist of the carboxy-terminal portion of nuc-1, bind specifically to upstream sequences of two of its target genes, pho2+ and pho-4+. these upstream sequenc ...19947969122
5-formyltetrahydropteroylpolyglutamates are the major folate derivatives in neurospora crassa conidiospores.5-formyltetrahydropteroylpolyglutamate (5-cho-h4pteglun) is the only reduced folate derivative that is stable to oxidation and alkaline ph. however, no metabolic role has been assigned to this folate derivative, and evidence for its existence in cells has been questioned. recently, serine hydroxymethyltransferase was shown to catalyze the formation of 5-cho-h4pteglun from 5,10-methenyl-h4pteglun (stover, p., and schirch, v. (1990) j. biol. chem. 265, 14227-14233). we have proposed that 5-cho-h4p ...19947961829
tertiary conformational changes of the neurospora crassa plasma membrane h(+)-atpase monitored by hydrogen/deuterium exchange kinetics. a fourier transformed infrared spectroscopy approach.attenuated total reflection fourier transform infrared spectroscopy of hydrated films of the neurospora crassa plasma membrane h(+)-atpase has been used to monitor the alpha-helix and beta-sheet contents and amide hydrogen exchange rates of the enzyme in the absence of ligands or locked in several stages of the enzyme catalytic cycle by mgadp, mg-vanadate, and mgatp-vanadate. no difference larger than 2% was found in the alpha-helix or beta-sheet content of the h(+)-atpase in different conformat ...19947961652
characterization of mitochondrial dna topoisomerase i from neurospora crassa.dna topoisomerase i isolated from the lower eukaryote neurospora crassa mitochondria was characterized. molar mass of the enzyme in the native state is 120 kda and 60-65 kda when denatured. the ph optimum of the enzyme is 7.8 and the kcl optimum concentration is 40 mmol/l. this topoisomerase is independent of atp and mg2+. n-ethylmaleimide, 4-chloromercuribenzoate, sds, guanidinium chloride, polyethylene glycol, heparin and ethidium bromide inhibit its activity, while novobiocin, nalidixic acid, ...19947959426
a cosmid with a hyr marker for fungal library construction and screening.the construction of a double-cos-site cosmid vector, pmocosx, for use in making filamentous fungal genomic dna libraries, is described. the vector has features that allow for selection of clones introduced into fungi by transformation and for efficient chromosome walking experiments. these features include (i) two cos sites allowing for easy construction of libraries without requiring size selection of insert dna; (ii) an xhoi site for insertion of sau3ai or mboi partially digested genomic dna i ...19947959044
enzyme inactivation related to a hyperoxidant state during conidiation of neurospora crassa.the conidiation process of neurospora crassa is characterized by three morphogenetic steps: hyphal adhesion, aerial hyphal formation, and production of conidia. previous data indicated the occurrence of a hyperoxidant state at the onset of all three morphogenetic steps. because glutamine synthetase (gs) and the biosynthetic glutamate dehydrogenase [gdh(nadp)] enzymes are susceptible to inactivation by reactive oxygen species, we followed these enzyme activities during conidiation and under diffe ...19947952190
characterization of chitin synthase from botrytis cinerea.chitin synthase in a microsomal preparation from botrytis cinerea had an apparent km for udp-n-acetylglucosamine of 2.0 mm while nikkomycin z and polyoxin d inhibited enzyme activity competitively with apparent ki values of approximately 0.1 microm and 6 microm respectively. the organophosphorus fungicide edifenphos was a non-competitive inhibitor (ki(app) 54 microm). preincubation of microsomes for 2 h at 25 degrees c resulted in a maximum twofold stimulation of chitin synthase activity while p ...19947952170
the neurospora crassa chs-2 gene encodes a non-essential chitin synthase.chitin is a structural component of morphologically distinct structures assembled during various phases of growth and development in filamentous fungi. in neurospora crassa, at least three different dna fragments related to chitin synthase have been identified. in this study we cloned, sequenced and characterized the chitin synthase 2 structural gene (designated chs-2). the amino acid sequence deduced from the cloned chs-2 genomic dna fragments is very similar to that of chitin synthase genes is ...19947952169
temperature effects on the resetting of the phase of the neurospora circadian rhythm.various temperatures relative to a 25 degrees c control have been applied as phase-resetting agents in release-assay experiments using the conidiation rhythm of the mold neurospora crassa. the larger the difference in temperature from the 25 degrees c control, the stronger the phase-resetting effects. phase-resetting curves of the weak type (type 1) are observed for temperatures up to 28 degrees c and down to 22 degrees c, whereas temperatures above 28 degrees c and less than 22 degrees c genera ...19947949309
tissue-specific differences of the mitochondrial protein import machinery: in vitro import, processing and degradation of the pre-f1 beta subunit of the atp synthase in spinach leaf and root this study we report the first comparison of the mitochondrial protein import and processing events in two different tissues from the same organism. both spinach leaf and root mitochondria were able to import and process the in vitro transcribed and translated neurospora crassa f1 beta subunit of atp synthase to the mature size product. temperature optimum for protein import, 20 degrees c, was considerably lower than that found in other systems. in spinach leaf mitochondria, the processing pe ...19947948913
(1,3) beta-glucan synthase activity of neurospora crassa: identification of a substrate-binding protein.(1,3)beta-glucan synthase activity from the filamentous ascomycete neurospora crassa was purified 1300-fold to a specific activity of 14,000 nmol glucose incorporated/min per mg protein. hyphae were disrupted and crude membrane fractions obtained by high-speed centrifugation. membrane fractions were extracted with tergitol np-40 and a second high-speed particulate fraction was obtained. enzyme activity was solubilized with (3-((3-cholamidopropyl)dimethylammonio)-1-propanesulfonate and octyl-beta ...19947947947
complementary dna sequences of the 24 kda and 21 kda subunits of complex i from neurospora.we have cloned and sequenced cdnas coding for two subunits of the peripheral arm of neurospora crassa complex i. the two polypeptides are synthesized as precursor proteins which are processed to mature forms with predicted molecular masses of 24331 and 20982 da.19947947902
substrate analogs as mechanistic probes for the bifunctional chorismate synthase from neurospora crassa.analogs of epsp (4-8) have been prepared, and their activity as substrates for the chorismate synthase from neurospora crassa has been characterized kinetically. the enzyme appears to show strict discrimination against substitution at the z-position of the enol ether side chain as well as against substitution at the s-position of the reduced analogs. both the glycolyl and (r)-lactyl analogs 4 and (r)-5 are good substrates, with (r)-5 having a higher v value than the natural substrate. three subs ...19947947820
nadh: ubiquinone oxidoreductase in obligate aerobic yeasts.the strictly aerobic yeasts candida pinus, cryptococcus albidus, rhodotorula minuta, rhodotorula mucilaginosa and trichosporon beigelii possess mitochondrial nadh dehydrogenases with significant features of the nadh:ubiquinone oxidoreductase (complex i). these species show in all growth phases and under standard cultivation conditions, nadh dehydrogenases of approximately 700 kda, which are sensitive to rotenone, a specific inhibitor of this complex. identical results were obtained with the weak ...19947941733
dna photolyase from the fungus neurospora crassa. purification, characterization and comparison with other photolyases.a phr-gene from the filamentous fungus neurospora crassa was overexpressed in escherichia coli cells, yielding a biologically active photolyase. after purification till apparent homogeneity, the 66 kda protein was found to contain equimolar amounts of 5,10-methenyltetrahydrofolic acid (mthf) and fad, classifying it as an mthf-type photolyase. compared to other mthf photolyases the absorption maximum of neurospora photolyase is shifted from ca 380 nm to 391 nm (epsilon = 34,800), while an additio ...19947938208
cloning and characterization of a neurospora crassa gene required for (1,3) beta-glucan synthase activity and cell wall formation.the glucan synthase 1 gene (gs-1) is required for (1,3) beta-glucan synthase activity [e.c.; udp glucose:1,3-beta-d-glucan 3-beta-d-glucosyltransferase] and for cell wall formation. the gs-1 gene was cloned by functional complementation of the cell-wall-less defect of the (1,3) beta-glucan synthase-deficient mutant, tm1, by using a genomic neurospora crassa cosmid library. a 2568-nucleotide gs-1 cdna sequence revealed a 532-amino acid open reading frame encoding a polypeptide of 59 kda. ...19947937796
a targeted-replacement system for identification of signals for de novo methylation in neurospora crassa.transformation of eukaryotic cells can be used to test potential signals for dna methylation. this approach is not always reliable, however, because of chromosomal position effects and because integration of multiple and/or rearranged copies of transforming dna can influence dna methylation. we developed a robust system to evaluate the potential of dna fragments to function as signals for de novo methylation in neurospora crassa. the requirements of the system were (i) a location in the n. crass ...19947935421
mitochondrial grpe is present in a complex with hsp70 and preproteins in transit across membranes.we characterized a 24-kda protein associated with matrix hsp70 (mt-hsp70) of neurospora crassa and saccharomyces cerevisiae mitochondria. by using specific antibodies, the protein was identified as mge, a mitochondrial homolog of the prokaryotic heat shock protein grpe. mge extracted from mitochondria was quantitatively bound to hsp70. it was efficiently released from hsp70 by the addition of mg-atp but not by nonhydrolyzable atp analogs or high salt. a mutant mt-hsp70, which was impaired in rel ...19947935381
cytoplasmic dynein and actin-related protein arp1 are required for normal nuclear distribution in filamentous fungi.cytoplasmic dynein is a multisubunit, microtubule-dependent mechanochemical enzyme that has been proposed to function in a variety of intracellular movements, including minus-end-directed transport of organelles. dynein-mediated vesicle transport is stimulated in vitro by addition of the glued/dynactin complex raising the possibility that these two complexes interact in vivo. we report here that a class of phenotypically identical mutants of the filamentous fungus neurospora crassa are defective ...19947929559
internal translational initiation in the mrna from the neurospora crassa albino-3 gene.the "ribosome scanning model" for translational initiation predicts that eukaryotic mrnas should, as a rule, be monocistronic. however, cases have recently been described of eukaryotic mrnas producing more than one protein through alternative translational initiation at several different aug codons. the present work reports the occurrence of multiple translational start sites on the mrna of the neurospora crassa gene albino-3 (al-3), encoding the carotenoid biosynthetic enzyme geranylgeranyl-pyr ...19947929398
a high throughput in vitro assay for fungal (1,3)beta-glucan synthase in vitro assay for (1,3)beta-glucan synthase activity from the filamentous ascomycete neurospora crassa, suitable for use as a high throughput screen for enzyme inhibitors is described. samples were added to 25 microliters reaction mixtures in 96-well v-bottom microtiter plates and plates incubated at 22 degrees c. reactions were terminated by the addition of tca and the contents transferred to a milliblot d apparatus containing inotech 201-a glass fiber filters. filters were washed to remove ...19947928688
isolation and characterization of cdna and genomic promoter region for a heat shock protein 30 from aspergillus nidulans.a cdna encoding for a heat shock protein 30 (hsp30) of aspergillus nidulans and the promoter region of its gene were analyzed for their primary structures. the promoter region had no heat shock element but possessed three inverted repeat sequences. northern blot hybridization indicated that the expression of the hsp30 gene was high at a normal temperature and was slightly accelerated at elevated temperatures in a. nidulans cells. although the deduced amino acid sequence of the a. nidulans hsp30 ...19947918658
dna uptake stimulating protein from neurospora crassa enhances dna and oligonucleotide uptake also in mammalian cells.the presence of the dna uptake stimulating protein (designated earlier as dusf by schablik and szabó (1981) fems microbiol. lett. 10, 395-397) was demonstrated on the surface of neurospora crassa (fgsc 1118, slime) fungal cells as well as on hep-2 and k562 human and f4n mouse tumor cells by immunofluorescence microscopy. dusf markedly enhanced the uptake of both macromolecular [3h]dna and of [3h]oligonucleotides by k562, hl-60 and also by dd human leukemia cells. polyclonal anti-dusf antibodies ...19947918627
cloning and sequence analysis of an h(+)-atpase-encoding gene from the human dimorphic pathogen histoplasma capsulatum.a gene related to the pma1 gene from saccharomyces cerevisiae was isolated from the pathogenic human dimorphic fungus, histoplasma capsulatum, using fungal-specific oligodeoxyribonucleotide (oligo) probes. this gene has been given the name hc-pma1. the structural organization of hc-pma1 consists of three exons (375, 2329 and 44 bp) and two introns (115 and 116 bp). the nucleotide sequence predicts an h(+)-atpase-related protein of 916 amino acids (aa). comparison of the deduced aa sequence to th ...19937916725
structure and sequence of the calmodulin gene from neurospora crassa.cdna and genomic clones of neurospora calmodulin were obtained by pcr. characterization revealed an open reading frame encoding a predicted protein of 149 amino acids, showing 85% identity to the human calmodulin protein sequence. comparison of the cdna and genomic sequence reveals the position of five introns, organized differently than is found in calmodulin genes from higher eukaryotes.19937916628
correlation of the physical and genetic maps of the centromeric region of the right arm of linkage group iii of neurospora crassa.we have cloned three linked genes serine-1 (ser-1), proline-1 (pro-1) and acetate-2 (ace-2) that lie near the centromere on the right arm of linkage group iii (lgiiir) of neurospora crassa. the ser-1 gene was cloned by sib selection. a chromosomal walk that spans 205 kilobases (kb) was initiated from ser-1. complementation analysis with clones isolated during the walk allowed identification of the pro-1 and ace-2 genes. restriction fragment length polymorphism analysis has confirmed the localiza ...19947912215
a polyprotein precursor of two mitochondrial enzymes in neurospora crassa. gene structure and precursor processing.n-acetylglutamate kinase (agk) and n-acetyl-gamma-glutamyl-phosphate reductase (agpr) function as two separate mitochondrial enzymes, but are encoded by a single nuclear gene in several fungi. the neurospora crassa arg-6 gene encoding these enzymes has been cloned and sequenced, and the enzymes responsible for processing the polyprotein precursor have been identified. the 871-amino acid precursor contains a normal n-terminal mitochondrial targeting sequence, an internal connecting region (approx ...19947907589
regulation of ammonium ion assimilation enzymes in neurospora crassa nit-2 and ms-5 mutant neurospora crassa the nit-2 and nmr-1 (ms-5) loci represent the major control genes encoding regulatory proteins that allow the coordinated expression of various systems involved with the utilization of a secondary nitrogen source. in this paper we examined the effect of the nit-2 and ms-5 (nmr-1 locus) mutations on the regulation of the ammonium assimilation enzymes, glutamine synthetase and glutamate dehydrogenase, which are regulated by the products of these genes; however, glutamate synth ...19937907211
cloning and characterization of centromeric dna from neurospora crassa.the centromere locus from linkage group vii of neurospora crassa has been cloned, characterized, and physically mapped. the centromeric dna is contained within a 450-kb region that is recombination deficient, a+t-rich, and contains repetitive sequences. repetitive sequences from within this region hybridize to a family of repeats located at or near centromeres in all seven linkage groups of n. crassa. genomic southern blots and sequence analysis of these repeats revealed a unique centromere stru ...19947904723
characterization of the cit-1 gene from neurospora crassa encoding the mitochondrial form of citrate synthase.we have isolated the cdna and corresponding genomic dna encoding citrate synthase in neurospora crassa. analysis of the protein coding region of this gene, named cit-1, indicates that it specifies the mitochondrial form of citrate synthase. the predicted protein has 469 amino acids and a molecular mass of 52,002 da. the gene is interrupted by four introns. hybridization experiments show that a cit-1 probe binds to two different fragments of genomic dna, which are located on different chromosomes ...19947904043
the role of the nad-dependent glutamate dehydrogenase in restoring growth on glucose of a saccharomyces cerevisiae phosphoglucose isomerase mutant.phosphoglucose isomerase pgi1-deletion mutants of saccharomyces cerevisiae cannot grow on glucose as the sole carbon source and are even inhibited by glucose. these growth defects could be suppressed by an over-expression on a multi-copy plasmid of the structural gene gdh2 coding for the nad-dependent glutamate dehydrogenase. gdh2 codes for a protein with 1092 amino acids which is located on chromosome xii and shows high sequence similarity to the neurospora crassa nad-glutamate dehydrogenase. s ...19937901008
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
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
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
high frequency repeat-induced point mutation (rip) is not associated with efficient recombination in neurospora.duplicated dna sequences in neurospora crassa are efficiently detected and mutated during the sexual cycle by a process named repeat-induced point mutation (rip). linked, direct duplications have previously been shown to undergo both rip and deletion at high frequency during premeiosis, suggesting a relationship between rip and homologous recombination. we have investigated the relationship between rip and recombination for an unlinked duplication and for both inverted and direct, linked duplica ...19947896093
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
two nuclear-coded subunits of mitochondrial complex i are similar to different domains of a bacterial formate hydrogenlyase subunit.a computer comparison of protein sequences revealed similarity between the 30.4 kda subunit of complex i from the fungus neurospora crassa and the orf5 subunit of formate hydrogenlyase from escherichia coli. the orf5 protein was previously known to be homologous to the 49 kda component of the mitochondrial enzyme. we show that the 30.4 kda corresponds to the n-terminal part while the 49 kda subunit corresponds to the c-terminal portion of the bacterial protein. thus, this bacterial protein repre ...19947890119
a phosphate-repressible, high-affinity phosphate permease is encoded by the pho-5+ gene of neurospora crassa.the pho-5+ gene of neurospora crassa, which encodes a high-affinity phosphate permease, has been cloned and analyzed. the deduced orf of 1707 nucleotides is interrupted by a single 63-nt intron and codes for a protein of 569 amino acids (aa). this aa sequence has 48% identity with the high-affinity phosphate transporter of saccharomyces cerevisiae, pho84. the pho-5 null mutants have no obvious phenotype. strains which contain a null mutation in pho-4, which encodes an additional high-affinity ph ...19957883177
mutations in the structural gene for cytochrome c result in deficiency of both cytochromes aa3 and c in neurospora crassa.the cyt-12-12 mutant of neurospora crassa is characterized by slow growth and a deficiency of spectrophotometrically-detectable cytochromes aa3 and c. using a sib-selection procedure we have isolated the cyt-12+ allele from a cosmid library of n. crassa genomic dna. characterization of the cyt-12+ allele reveals that it encodes the structural gene for cytochrome c. dna sequence analysis of the cyt-12-12 allele revealed a mutation in the cytochrome c coding sequence that results in replacement of ...19947882427
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
disruption of the cyclosporin synthetase gene of tolypocladium niveum.cyclosporin a is a potent and clinically-important immunosuppressive drug (sandimmunr). it is produced by the fungus tolypocladium niveum. a transformation system for t. niveum atcc34921 based on hygromycin selection was established. in order to obtain a t. niveum promoter, the cyclophilin gene was isolated using the neurospora crassa gene as probe. a plasmid vector was constructed in which the promoter region of the t. niveum cyclophilin gene was fused to a bacterial hygromycin phosphotransfera ...19947874740
the function and specificity of the c-terminal tripeptide glyoxysomal targeting signal in neurospora crassa.the function of the c-terminal tripeptide targeting signal responsible for microbody targeting in many eukaryotes has been investigated in the filamentous fungus neurospora crassa. using an in-vivo targeting assay that employs transformants carrying c-terminally-modified versions of the bacterial enzyme chloramphenicol acetyltransferase (cat), it has been demonstrated that c-terminal tripeptide-dependent import occurs most efficiently in response to nutritional acetate-induction. under these con ...19947874736
triethylenemelamine: induction of specific-locus mutations in the ad-3 region of heterokaryon 12 of neurospora crassa.the mutagenicity of the trifunctional alkylating (or cross-linking) agent tem (triethylenemelamine or 2,4,6-tris(1-aziridinyl)-1,3,5-triazine) in the adenine-3 (ad-3) region was studied with a two-component heterokaryon (h-12) of neurospora crassa. the objective was to characterize the genetic damage produced by this chemical to determine the spectrum of specific-locus mutations induced in a lower eukaryotic organism and to compare this spectrum with that induced in the mouse. specific-locus mut ...19957870102
neurospora crassa blue light-inducible gene light induces various physiological, morphological and biochemical reactions in the filamentous fungus neurospora crassa. this light response is accompanied by a global change in gene expression, and several light-inducible transcripts (bli-genes) have been cloned. we isolated the genomic clone of the gene bli-3, whose mrna we have previously shown to be induced 2 minutes after the beginning of illumination. its dna sequence predicted a transcriptional unit of 1050 bp encoding a novel, hydr ...19947866300
the neurospora transposon tad is sensitive to repeat-induced point mutation (rip).rip (repeat-induced point mutation) efficiently mutates repeated sequences in the sexual phase of the neurospora crassa life cycle. nevertheless, an active line-like retrotransposon, tad, was found in a n. crassa strain from adiopodoumé. the possibility was tested that tad might be resistant to rip, or that the adiopodoumé strain might be incompetent for rip. tad elements derived from the adiopodoumé strain were found to be susceptible to rip. in addition, strains lacking active tad elements, in ...19947851763
neurospora mutants affecting polyamine-dependent processes and basic amino acid transport mutants resistant to the polyamine inhibitor, alpha-difluoromethylornithine.polyamines (spermidine and spermine) are required by living cells, but their functions are poorly understood. mutants of neurospora crassa with enhanced or diminished sensitivity to interference with polyamine synthesis, originally selected to study the regulation of the pathway, were found to have unexpected defects. a group of four non-allelic mutations, causing no interference with polyamine synthesis, each imparted spermidine auxotrophy to a genotype already partially impaired in spermidine ...19947851762
light and developmental regulation of the gene con-10 of neurospora crassa.the gene con-10 of neurospora crassa is expressed preferentially during conidiation and following illumination of vegetative mycelia with blue light. in this study we have examined the segmental locations of the genetic elements associated with con-10 that are responsible for light and developmental expression. a translational fusion was prepared between the initial segment of con-10 and escherichia coli lacz. deletions were then introduced into the con-10 upstream region associated with this tr ...19957851642
suppression of gene expression by homologous transgenes.when a wild-type strain of neurospora crassa is transformed with different portions of the carotenogenic albino 1 or albino 3 genes, up to 30-35% of the transformants show an albino phenotype. the albino transformants presented a variety of phenotypes ranging from white or yellow to dark yellow colour. the ectopically integrated sequences provoke a severe impairment of the expression of the endogenous al-1 or al-3 genes. this phenomenon, that has been termed quelling, is found to be spontaneousl ...19947847887
nitrogen regulation in fungi.nitrogen regulation has been extensively studied in fungi revealing a complex array of interacting regulatory genes. the general characterisation of the systems in aspergillus nidulans and neurospora crassa shall be briefly described, but much of this paper will concentrate specifically on the recent molecular characterisation of area, the principle regulatory gene from a. nidulans which mediates nitrogen metabolite repression. three areas shall be explored in detail, firstly the dna binding dom ...19947847882
characterization and expression of an antifungal zeamatin-like protein (zlp) gene from zea mays.a cdna clone encoding a basic thaumatin-like protein of zea mays was recovered from a mid-development seed cdna library. the gene, zlp, encoded a protein that was nearly identical with maize zeamatin and alpha-amylase/trypsin inhibitor. expression of zlp mrna was highest in the endosperm tissue of seed 4 weeks after pollination. expression of zeamatin-like (zlp) protein correlated with mrna; also, a low basal level of zlp expression in leaf was not appreciably induced by abiotic stresses. zlp wa ...19947846159
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
endosomal accumulation of ph indicator dyes delivered as acetoxymethyl esters.intracellular distributions of the putative cytosolic ph indicator dyes bcecf [2',7'-bis-(2-carboxyethyl)-5(and 6)-carboxyfluorescein], c.snarf [5(and 6)-carboxy-seminaphthorhodafluor-1], and c.snarf-calcein have been examined in neurospora crassa and in murine fibroblasts (nih-3t3 cells) under conditions in which both kinds of cells produce visible microscopic vacuoles. all three dyes were administered in electroneutral forms, with the hydroxyl and carboxyl groups esterified (designated as -am ...19947823037
the saccharomyces cerevisiae homologue of ribosomal protein s26.the nucleotide sequence of rps26, the gene encoding a homologue of ribosomal protein small subunit s26 in saccharomyces cerevisiae, was determined. the deduced amino-acid sequence showed significant identity with its counterparts from neurospora crassa, human, rat and arabidopsis thaliana. disruption of rps26 resulted in the formation of micro-colonies, suggesting that it is important for the normal cell growth of s. cerevisiae.19947821815
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
molecular characterization of upstream regulatory sequences controlling the photoinduced expression of the albino-3 gene of neurospora the filamentous fungus neurospora crassa the biosynthesis of carotenoids is regulated by blue light, principally through transcriptional activation of some key genes in the carotenogenic enzymatic pathway. here we report the characterization of the photoinducible promoter of the albino-3 (al-3) gene, encoding ggpp synthase. we have modified the 5' non-coding sequence of the cloned al-3 gene by deletion and site-directed mutagenesis, and we have tested the residual photoinducibility of the dif ...19947815938
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
heat shock inhibits and activates different protein degradation pathways and proteinase activities in neurospora neurospora crassa, heat shock treatment inhibits proteolytic activity. atp-independent proteinases were analysed after polyacrylamide gel electrophoresis using renaturing gelatine gels. proteinases of 24, 29, and 130 kda were shown to be inhibited by heat shock and were further characterized as to their properties. a major part of the heat shock-induced inhibition is probably due to suppression of de novo synthesis of proteinases as deduced from experiments with cycloheximide. during several ...19947813891
dna binding site specificity of the neurospora global nitrogen regulatory protein nit2: analysis with mutated binding sites.nit2, a positive-acting regulatory protein in neurospora crassa, activates the expression of a series of unlinked structural genes that encode nitrogen catabolic enzymes. nit2 binding sites in the promoter regions of nit3, alc and lao have at least two gata sequence elements. we have examined the binding affinity of the nit2 protein for the yeast dal5 wild-type upstream activation sequence uasntr, which contains two gata elements, and for a series of mutated binding sites, each differing from th ...19947808401
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