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molecular cloning of a neurospora crassa carotenoid biosynthetic gene (albino-3) regulated by blue light and the products of the white collar genes.the albino-3 (al-3) gene of neurospora crassa, which probably encodes the carotenoid biosynthetic enzyme geranylgeranyl pyrophosphate synthetase, was cloned. the n. crassa triple mutant al-3 qa-2 aro-9 was transformed to qa-2+ with mixtures of plasmids bearing n. crassa dna inserts, and the transformants were screened for the al-3+ phenotype. one al-3+ qa-2+ transformant (al3-1) was examined in detail and shown to contain intact vector sequences integrated into the n. crassa genome. the vector a ...19892524647
photoaffinity labelling of mitochondrial nadh: ubiquinone reductase with pethidine analogues.1. chemically reactive derivatives of pethidine analogues--novel potent inhibitors of the mitochondrial nadh: ubiquinone reductase (complex i)--were synthesized. 2. dose-response curves of these components revealed that the photoactivatable aryl azido derivative has retained most of the inhibitory activity displayed by the parent substance. after introduction of a radioactive iodine isotope into the molecule, it was used as a probe for the localization of the inhibitor binding polypeptides withi ...19892525381
translocation arrest by reversible folding of a precursor protein imported into mitochondria. a means to quantitate translocation contact sites.passage of precursor proteins through translocation contact sites of mitochondria was investigated by studying the import of a fusion protein consisting of the nh2-terminal 167 amino acids of yeast cytochrome b2 precursor and the complete mouse dihydrofolate reductase. isolated mitochondria of neurospora crassa readily imported the fusion protein. in the presence of methotrexate import was halted and a stable intermediate spanning both mitochondrial membranes at translocation contact sites accum ...19892529262
oxidation of neurospora crassa nadp-specific glutamate dehydrogenase by activated oxygen species.the glutamine synthetase and the nadp-specific glutamate dehydrogenase activities of neurospora crassa were lost in a culture without carbon source only when in the presence of air. glutamine synthetase was previously reported to be liable to in vitro and in vivo inactivation by activated oxygen species. here we report that nadp-specific glutamate dehydrogenase was remarkably stable in the presence of activated oxygen species but was rendered susceptible to oxidative inactivation when chelated i ...19892530208
a unique low frequency raman band associated with metal binding to metallothionein.we find that the low frequency raman spectrum of zn(ii) metallothionein has a single prominent band at 138 cm-1 which is absent from the raman spectrum of the metal-free protein. this feature is also found for cd(ii) binding to both of the independent metallothionein domains and the metallothionein from neurospora crassa. tco(iii) coordination to metallothionein results in a similar raman band which is also found for the complex (ph4as)[reo(sch2ch2s)2]. by comparing these results to literature d ...19892528950
stereoselective arginine binding is a phylogenetically conserved property of group i self-splicing rnas.we have examined the reaction of gtp with rna polymerase transcripts containing the self-splicing rna precursors from the neurospora crassa cob1 intron, and from introns in the suny, nrdb and td genes of bacteriophage t4. in each case, we find a low km for gtp (between 0.8 and 11 microm), accompanied by competitive inhibition of the gtp reaction by l-arginine, as was found for the previously examined tetrahymena nuclear pre-rrna intron. trials with the 20 standard amino acids show that inhibitio ...19892531082
amino acid sequence of the alpha and beta subunits of methanosarcina barkeri atpase deduced from cloned genes. similarity to subunits of eukaryotic vacuolar and f0f1-atpases.the atpa and atpb genes coding for the alpha and beta subunits, respectively, of membrane atpase were cloned from a methanogen methanosarcina barkeri, and the amino acid sequences of the two subunits were deduced from the nucleotide sequences. the methanogenic alpha (578 amino acid residues) and beta (459 amino acid residues) subunits were highly homologous to the large and small subunits, respectively, of vacuolar h+-atpases; 52% of the residues of the methanogenic alpha subunit were identical ...19892544575
inhibition by aluminum hydroxide of the voltage-dependent closure of the mitochondrial channel, vdac.micromolar quantities of aluminum have been found (dill et al. (1987) j. membrane biol. 99, 187-196) to reduce the voltage dependence of the mitochondrial outer membrane channel, vdac, from neurospora crassa. in the present study, various metallic and organic ions were tested for possible aluminum-like effect, and only the trivalent metals exhibited a similar ability to reduce the channels voltage dependence. however, trivalency alone was not sufficient because lanthanum (iii) had no effect. qua ...19892469483
a mitochondrial protein from neurospora crassa detected both on ribosomes and in membrane fractions. analysis of the gene, the message, and the protein.we have isolated clones representing at least three nuclear genes for mitochondrial ribosomal proteins from neurospora crassa by screening a lambda gt11 cdna library with an antiserum against a mixture of these proteins. the cdna and genomic dna sequence for one of these genes, mrp-3, was determined. the mrp3 protein was purified by immune-affinity chromatography, using a monoclonal antibody probe, and subjected to amino acid sequence analysis to identify the mature amino terminus and a prospect ...19892521217
pneumocystis carinii: sequence from ribosomal rna implies a close relationship with fungi.pneumocystis carinii is the etiologic agent of a lethal pneumonia which occurs in patients with the acquired immune deficiency syndrome (aids) and other immunocompromised hosts. the basic biochemical and genetic characteristics of p. carinii are poorly understood and its taxonomic classification as a protozoan is uncertain. to address the taxonomic question, a method was developed for the extraction of total rna from p. carinii. denaturing agarose gel electrophoresis showed the two ribosomal rna ...19892470612
nucleotide sequence of the neurospora crassa trp-3 gene encoding tryptophan synthetase and comparison of the trp-3 polypeptide with its homologs in saccharomyces cerevisiae and escherichia coli.the complete nucleotide sequence of the neurospora crassa trp-3 gene-encoding tryptophan synthetase has been determined; we present an analysis of its structure. a comparison of the deduced amino acid sequence of the trp-3 polypeptide with its homologs in saccharomyces cerevisiae (encoded by the trp5 gene) and escherichia coli (encoded by the trpa and trpb genes) shows that the a and b domains (amino acid segments homologous to the trpa and trpb polypeptides, respectively) of the n. crassa and y ...19892521855
[properties of 2,5-diamino-4-oxy-6-ribosylaminopyrimidine-5'- phosphate reductase, a enzyme of the second stage of flavinogenesis in pichia guilliermondii yeasts].2,5-diamino-4-oxy-6-ribosylaminopyrimidine-5'-phosphate reductase has been isolated from cells of pichia guilliermondii and subjected to 20-fold purification by treating extracts with streptomycin sulphate, frationating proteins (nh4)2so4 at 45-75% of saturation and chromatography on blue sepharose cl-6b. the use of gel filtration through sephadex g-150 and chromatography on deae-cellulose proved to be less effective for the enzyme purification. it has been established that it is 2,5-diamino-4-o ...19892511652
evidence for three differentially regulated catalase genes in neurospora crassa: effects of oxidative stress, heat shock, and development.genetic and biochemical studies demonstrated that neurospora crassa possesses three catalases encoded by three separate structural genes. the specific activities of the three enzymes varied in response to superoxide-mediated stress, heat shock, and development. the three loci, which we designated cat-1, cat-2, and cat-3, map to the right arms of chromosomes iii, vii, and iii, respectively. the cat-1-encoded enzyme (designated cat-1; estimated molecular weight, 315,000; pi 5.2) was the predominan ...19892540152
x-ray-induced specific locus mutations in the ad-3 region of two-component heterokaryons of neurospora crassa. iii. genetic fine structure analysis of the ad-3 and immediately adjacent genetic regions by means of complementation tests.genetic fine structure analysis of the ad-3 and immediately adjacent genetic regions was made by means of complementation tests on all possible pairwise combinations of 50 x-ray-induced irreparable adenine-3 mutants (designated ad-3ir). all mutants were induced in either heterokaryon 11 or heterokaryon 12 of neurospora crassa, 2-component heterokaryons heterozygous for mutants at the 3 closely linked loci ad-3a and ad-3b and nic-2 (nicotinamide-requiring) located about 5.0 map units distal to ad ...19892522164
molybdenum cofactor biosynthesis in humans. identification of two complementation groups of cofactor-deficient patients and preliminary characterization of a diffusible molybdopterin precursor.molybdenum cofactor deficiency is a devastating disease with affected patients displaying the symptoms of a combined deficiency of sulfite oxidase and xanthine dehydrogenase. because of the extreme lability of the isolated, functional molybdenum cofactor, direct cofactor replacement therapy is not feasible, and a search for stable biosynthetic intermediates was undertaken. from studies of cocultured fibroblasts from affected individuals, two complementation groups were identified. coculture of g ...19892522104
development of an in vitro transcription system for neurospora crassa mitochondrial dna and identification of transcription initiation sites.we have developed an in vitro transcription system for neurospora crassa mitochondrial dna (mtdna) and used it to identify transcription initiation sites at the 5' ends of the genes encoding the mitochondrial small and large rrna and cytochrome b (cob). the in vitro transcription start sites correspond to previously mapped 5' ends of major in vivo transcripts of these genes. sequences around the three transcription initiation sites define a 15-nucleotide consensus sequence, 5'-ttagara(t/g)g(t/g) ...19892528684
pneumocystis carinii: taxing taxonomy.the detailed taxonomy of pneumocystis carinii has not been decided. most experts agree that it is either a fungus or a protozoan. in favor of a fungal designation are the recent reports that 16s and 18s rrna sequences of p. carinii have greater homology to saccharomyces cerevisiae and neurospora crassa than to selected protozoa. other studies show that sequences of the 18s and 26s rrna, microtubular ultrastructure, membrane fusion potential and dna content per cell resemble protozoa more than fu ...19892477272
transformation of neurospora crassa by an integrative transforming plasmid is not enhanced by ribosomal dna sequences.two integrative transforming plasmids of neurospora crassa that differed only by the presence of almost all of a ribosomal dna repeat unit on one plasmid were constructed. the plasmids were used to test the target concentration hypothesis which states that the transformation frequency is proportional to the number of genomic copies of a homologous sequence located on the transforming plasmid. since there are approx. 200 copies of the rdna sequences in the genome, the target concentration hypothe ...19892525404
molecular cloning and expression in saccharomyces cerevisiae and neurospora crassa of the invertase gene from neurospora crassa.a plasmid (named pcn2) carrying a 7.6 kb bamhi dna insert was isolated from a neurospora crassa genomic library raised in the yeast vector yrp7. saccharomyces cerevisiae suco and n. crassa inv strains transformed with pnc2 were able to grow on sucrose-based media and expressed invertase activity. saccharomyces cerevisiae suco (pnc2) expressed a product which immunoreacted with antibody raised against purified invertase from wild type n. crassa, although s. cerevisiae suc+ did not. the cloned dna ...19892531138
bialaphos resistance as a dominant selectable marker in neurospora crassa.conidia of neurospora crassa are sensitive to the herbicide bialaphos at concentrations of 160 mg/l in westergaard's or fries' minimal media. plasmid pja4 was constructed by inserting a truncated bar gene from streptomyces hygroscopicus fused to the his-3 promoter from n. crassa into puc19. the bar gene in plasmid pja4 confers resistance to bialaphos when transformants are selected on a medium containing bialaphos. the bar gene can be used as an additional dominant selectable marker for transfor ...19892532965
yeast cyclophilin: isolation and characterization of the protein, cdna and gene.cyclophilin (cph) has been isolated from the yeast saccharomyces cerevisiae, purified to homogeneity and partially sequenced. oligodeoxyribonucleotides deduced from this sequence were used to isolate the corresponding cdna and gene. an open reading frame coding for a 162-amino acid (aa) protein with a calculated mr of 17,392, was deduced from the nucleotide sequence. comparison between yeast and human cph shows a very high overall sequence conservation (65% aa homology). the binding of yeast cph ...19892687115
improved transformation efficiency of aspergillus niger using the homologous niad gene for nitrate reductase.aspergillus niger transformation frequencies of up to 1,176 transformants per micrograms dna were achieved using the plasmid vector psta10 containing the a. niger nitrate reductase structural gene. analysis of genomic endonuclease cleaved dna from nitrate utilising transformants by dna hybridisation, showed that most integration events are as a result of homologous recombination. the niad transformation system was used successfully for the introduction of the unselected escherichia coli fusion g ...19892791035
transformation by integration in podospora anserina. iii. replacement of a chromosome segment by a two-step process.we have developed in podospora anserina a two-step procedure for dna sequence replacement through transformation which might be applicable to other filamentous fungi. targeting of transforming dnas to their homologous locus is achieved provided a cosmid vector is used. southern blot analysis of genomic dnas from a set of transformants is presented. the data confirm that cosmids integrate into the chromosome through mostly homologous recombination which leads to a duplicated sequence separated by ...19892575706
two proteins encoded at the chla locus constitute the converting factor of escherichia coli chla1.molybdopterin (mpt) is not produced by the escherichia coli mutants chla1, chlm, or chln or by the neurospora crassa mutant nit-1. extracts of e. coli chla1 contain an activity, the converting factor, which is functionally defined by its ability to convert a low-molecular-weight precursor present in crude extracts of n. crassa nit-1 into molybdopterin in vitro. in this study, it has been shown that the converting factor consists of two associative proteins (10 and 25 kilodaltons [kda]) which can ...19892656653
molecular characterization of trp1, a gene coding for tryptophan synthetase in the basidiomycete coprinus cinereus.we utilized a cloned gene (trp5) encoding tryptophan synthetase (tsase) from saccharomyces cerevisiae to identify and clone the corresponding gene (trp1) from the basidiomycete coprinus cinereus. the primary nucleotide (nt) sequence of this gene was determined and compared to sequences from other filamentous fungi, as well as to other genes coding for tsase. a transformation assay was used to demonstrate that 321 nt, which do not include caat or tataaa elements and precede the translation initia ...19892806911
regulation of the nuclear genes encoding the cytoplasmic and mitochondrial leucyl-trna synthetases of neurospora crassa.we show that the nuclear genes for the cytoplasmic and mitochondrial leucyl-trna synthetase (leurs) of neurospora crassa are distinct in their encoded proteins, codon usage, mrna levels, and regulation. the 4.2-kilobase-pair region representing the structural gene for cytoplasmic leurs and flanking regions has been sequenced. the positions of the 5' and 3' ends of mrna and of a single 62-base-pair intron have been mapped. the methionine-initiated open reading frame encoded a protein of 1,123 ami ...19892532300
x-ray-induced specific-locus mutations in the ad-3 region of two-component heterokaryons of neurospora crassa. iv. irreparable mutants of genotype ad-3a and ad-3b result from multilocus deletion and an unexpectedly high frequency of multiple-locus mutations.the induction of specific-locus mutations in the ad-3 region of neurospora crassa after x-irradiation was studied in a two-component heterokaryon to determine: (1) the ratio of reparable ad-3 mutants (presumed gene/point mutations, designated ad-3r) to irreparable ad-3 mutants (presumed multilocus deletions, designated ad-3ir), and (2) the induction kinetics of each class (webber and de serres, 1965). more extensive genetic tests made subsequently (de serres, 1989a) on the 832 x-ray-induced spec ...19892529438
cytochrome oxidase subunit v gene of neurospora crassa: dna sequences, chromosomal mapping, and evidence that the cya-4 locus specifies the structural gene for subunit v.the sequences of cdna and genomic dna clones for neurospora cytochrome oxidase subunit v show that the protein is synthesized as a 171-amino-acid precursor containing a 27-amino-acid n-terminal extension. the subunit v protein sequence is 34% identical to that of saccharomyces cerevisiae subunit v; these proteins, as well as the corresponding bovine subunit, subunit iv, contain a single hydrophobic domain which most likely spans the inner mitochondrial membrane. the neurospora crassa subunit v g ...19892540423
sensitivity to cyclosporin a is mediated by cyclophilin in neurospora crassa and saccharomyces cerevisiae.cyclosporin a, a cyclic fungal undecapeptide produced by tolypocladium inflatum, is a potent immunosuppressive drug originally isolated as an antifungal antibiotic. cyclosporin a (csa) is widely used in humans to prevent rejection of transplanted organs such as kidney, heart, bone marrow and liver. the biochemical basis of csa action is not known: its primary cellular target has been suggested to be calmodulin, the prolactin receptor or cyclophilin, a csa-binding protein originally isolated from ...19892531848
nucleotide sequence and regulation of expression of the aspergillus nidulans gdha gene encoding nadp dependent glutamate dehydrogenase.the nucleotide sequence of the aspergillus nidulans gdha gene encoding nadp linked glutamate dehydrogenase has been determined and northern blot analysis used to study the regulation of expression of this gene. the gdha gene is 1485 nucleotides long and, by comparison with the corresponding neurospora crassa am gene, has two putative introns of 53 nucleotides and a protein encoding region of 1380 nucleotides that codes for an inferred protein of 49.63 kda which shows regions of homology with glu ...19892550758
reconstitution of a light-stimulated adenylate cyclase from retina and neurospora crassa preparations. characterization of the heterologous systems using normal and degenerative retinas.adenylate cyclase catalytic subunits from neurospora crassa membranes may interact with regulatory factors from membranes of bovine retinal rod outer segments (pretreated with n-ethylmaleimide), reconstituting a heterologous system which, in the presence of light, is catalytically active in assay mixtures containing mgatp. maximal activation was observed at 550 nm. transducin-depleted retinal membranes were not capable of reconstituting the heterologous light-stimulated adenylate cyclase system. ...19892553402
[identification of human porins. ii. characterization and primary structure of a 31-lda porin from human b lymphocytes (porin 31hl)].we characterize and describe for the first time the primary structure of a human porin with the molecular mass of 31 kda derived from the plasmalemm of b-lymphocytes (porin 31hl). porin 31hl is shown to be a basic, channel forming membrane protein. the protein chain is composed of 282 amino acids with a relative molecular mass of 30641 da without derivatisation. it is not a glycoprotein. the n-terminus is acetylated. altogether the amino-acid sequence shows 56% hydrophilic or charged amino acids ...19892559745
transformation in fungi.transformation with exogenous deoxyribonucleic acid (dna) now appears to be possible with all fungal species, or at least all that can be grown in culture. this field of research is at present dominated by saccharomyces cerevisiae and two filamentous members of the class ascomycetes, aspergillus nidulans and neurospora crassa, with substantial contributions also from fission yeast (schizosaccharomyces pombe) and another filamentous member of the class ascomycetes, podospora anserina. however, tr ...19892651864
cloning and analysis of the neurospora crassa gene for cytochrome c heme lyase.the cyt-2-1 mutant of neurospora crassa is deficient in cytochromes aa3 and c and in cytochrome c heme lyase activity (mitchell, m.b., mitchell, h.k., and tissieres, a. (1953) proc. natl. acad. sci. u.s.a. 39, 606-613; nargang, f.e., drygas, m.e., kwong, p.l., nicholson, d.w., and neupert, w. (1988) j. biol. chem. 263, 9388-9394). by rescue of the slow growth character of the cyt-2-1 mutant, we have cloned the cyt-2+ gene from a n. crassa genomic library using sib selection. analysis of the dna ...19892572587
acyl carrier protein is present in the mitochondria of plants and eucaryotic micro-organisms.proteins antigenically similar to the acyl carrier protein (acp) found in the mitochondria of neurospora crassa were detected by immunoblotting and radioimmunoassay techniques in mitochondria isolated from yeast, potatoes, and pea leaves. these mitochondrial proteins were similar to neurospora acp both in their electrophoretic mobility and in their unusual decrease in mobility upon reduction. authentic acp(s) show this type of change upon conversion of the acylated to the unacylated form. purifi ...19892680483
primary structure of cucumber (cucumis sativus) ascorbate oxidase deduced from cdna sequence: homology with blue copper proteins and tissue-specific expression.cdna clones for ascorbate oxidase were isolated from a cdna library made from cucumber (cucumis sativus) fruit mrna. the library was screened with synthetic oligonucleotides that encode the nh2-terminal sequence of this enzyme. nucleotide sequence analysis of the cloned cdna inserts revealed a 1761-base-pair open reading frame that encoded an nh2-terminal signal peptide of 33 amino acids and a mature enzyme of 554 amino acids (mr, 62,258). the amino acid sequence deduced from nucleotide sequence ...19892919172
dna sequence and secondary structures of the large subunit rrna coding regions and its two class i introns of mitochondrial dna from podospora anserina.dna sequence analysis has shown that the gene coding for the mitochondrial (mt) large subunit ribosomal rna (rrna) from podospora anserina is interrupted by two class i introns. the coding region for the large subunit rrna itself is 3715 bp and the two introns are 1544 (r1) and 2404 (r2) bp in length. secondary structure models for the large subunit rrna were constructed and compared with the equivalent structure from escherichia coli 23s rrna. the two structures were remarkably similar despite ...19892494353
genetic transformation system for the aflatoxin-producing fungus aspergillus flavus.a heterologous transformation system was developed for aspergillus flavus with efficiencies greater than 20 stable transformants per micrograms of dna. protoplasts of uracil-requiring strains of the fungus were transformed with plasmid and cosmid vectors containing the pyr-4 gene of neurospora crassa. transformants were selected for their ability to grow and sporulate on medium lacking uracil. vector dna appeared to integrate randomly into the genome of a. flavus with a tendency for multiple, ta ...19892495764
a nuclear mutant of neurospora crassa lacking subunit 1 of cytochrome c oxidase. 1989210181
immunochemical analysis shows that an atp/adp-translocator is associated with the inner-envelope membranes of amyloplasts from acer pseudoplatanus l.pure preparations of intact amyloplasts and chloroplasts, free from mitochondrial contamination, were isolated from cultured cells of the white-wild and green-mutant lines of sycamore (acer pseudoplatanus l.), respectively. a specific rabbit antiserum against yeast mitochondrial cytochrome c(1) only cross-reacted with mitochondrial membranes from the white-wild sycamore cells. the outer and inner envelope-membranes of the two plastid-types were isolated and subsequently analyzed by sodium dodecy ...198916666656
genetic transformation of the biocontrol fungus gliocladium virens to benomyl resistance.methodology was developed to isolate and regenerate protoplasts from the biocontrol fungus gliocladium virens and to transform them to benomyl resistance with a neurospora crassa beta-tubulin gene. southern blots demonstrated that multiple copies of the vector integrated into the chromosomal dna of stable biotypes but not of abortive transformants. analysis of nuclear condition in vegetative and asexual structures demonstrated that no structure of g. virens is dependably uninucleate and thus pre ...199016348312
occurrence of nitrate reductase and molybdopterin in xanthomonas maltophilia.fifteen of 23 atcc strains and 2 of 9 clinical isolates of xanthomonas maltophilia, all of which grew aerobically on ammonia, but not nitrate, as a sole nitrogen source, reduced nitrate to nitrite. x. maltophilia failed to grow anaerobically on complex medium with or without nitrate, so it is considered an obligate aerobe. nitrate-reducing strains contained reduced methyl viologen nitrate reductase (mvh-nr) with specific activities ranging from 49.2 to 192 u mg of protein. strain atcc 17666 doub ...199016348378
dna-mediated genetic changes in neurospora crassa.evidence for genetic transformation in neurospora crassa is based on the observations that allo-dna has a specific effect in producing transformants which is abolished by dnaase treatment and that iso-dna is not effective in transformation. here, unambiguous evidence for genetic transformation is provided by transfer of a temperature-sensitive inositol requirement from a donor to a recipient strain. data provided also suggest the role of growth conditions and the involvement of a nuclease gene i ...1990159941
cloning of the pyr4 gene encoding orotidine-5'-phosphate decarboxylase in cephalosporium acremonium.we have cloned the cephalosporium acremonium pyr4 gene by cross-hybridization with the equivalent gene from neurospora crassa, the closest relative from which this gene is available. the c. acremonium pyr4 gene complements an e. coli pyrf mutant lacking orotidine-5'-phosphate decarboxylase (ompdecase), and most probably does not contain introns. maxicell analysis in e. coli shows that it encodes a 46 kda polypeptide. the c. acremonium ompdecase contains a highly conserved pentadecapeptide charac ...19902140299
wheat germ agglutinin in wheat seedling roots: induction by elicitors and fungi.treatment of wheat (triticum aestivum l.) seedlings with elicitors originating from either plant or fungal cell walls induces about a 2-fold increase of wheat germ agglutinin (wga) in the roots. while the wga content in roots of healthy plants normally decreases as a function of germination time, a transient accumulation of wga could be observed in plants challenged with different fungi, including rhizoctonia solani, fusarium culmorum, pythium ultimum and neurospora crassa. peak levels in challe ...199024226823
regulation of inorganic sulfate activation in filamentous fungi. allosteric inhibition of atp sulfurylase by 3'-phosphoadenosine-5'-phosphosulfate.atp sulfurylases from penicillium chrysogenum, penicillium duponti, aspergillus nidulans, and neurospora crassa are strongly inhibited by 3'-phosphoadenosine-5'-phosphosulfate (paps), the product of the second (adenosine-5'-phosphosulfate kinase-catalyzed) reaction in the two-step activation of inorganic sulfate. the v versus [paps] plots are sigmoidal. at physiological concentrations of mgatp (0.17-3 mm) and so4(2-) (0.4-10 mm), the [i]0.5 for paps inhibition of the p. chrysogenum enzyme is 35- ...19902162344
nucleotide sequence and nuclear protein binding of the two regulatory sequences upstream of the am (gdh) gene in neurospora.we have constructed a series of deletions in the 5' non-coding sequences of the cloned neurospora crassa am gene which specifies nadp specific glutamate dehydrogenase. all of the deletions begin at -4.4 kb with respect to the am transcription start site and extend for various distances toward the am gene. using vectors with a truncated fragment of the am gene, we introduced these deletions into the chromosome upstream of am by transformation. analysis of glutamate dehydrogenase expression in str ...19902164625
import into mitochondria of precursors containing hydrophobic passenger proteins: pretreatment of precursors with urea inhibits import.we have studied the import into isolated yeast mitochondria of three hydrophobic passenger proteins attached to the n-terminal cleavable presequence of mitochondrial atpase subunit 9 from neurospora crassa. one natural precursor (pn9) contained n. crassa subunit 9; two chimaeric precursors, n9l/y8-1 and n9l/y9-2, respectively contained yeast mitochondrial atpase subunits 8 and 9. in the absence of urea, pn9 and n9l/y8-1 are imported efficiently but n9l/y9-2 is not imported. after pretreatment of ...19902168755
plant and fungus calmodulins are polyubiquitinated at a single site in a ca2(+)-dependent manner.in plants ca2+ plays a crucial role as second messenger. thus calmodulin is one of the most important signal transducing molecules for metabolic regulation in plants. previously we showed that bovine testis calmodulin can be covalently coupled at one site to ubiquitin in a ca2(+)-dependent manner in the presence of atp/mg2+ by ubiquityl-calmodulin synthetase. since calmodulin from spinach has 13 amino acid sequence differences to bovine calmodulin - two of them in ca2(+)-binding loops - it was u ...19902172031
import of adp/atp carrier into mitochondria: two receptors act in parallel.we have identified the yeast homologue of neurospora crassa mom72, the mitochondrial import receptor for the adp/atp carrier (aac), by functional studies and by cdna sequencing. mitochondria of a yeast mutant in which the gene for mom72 was disrupted were impaired in specific binding and import of aac. unexpectedly, we found a residual, yet significant import of aac into mitochondria lacking mom72 that occurred via the receptor mom19. we conclude that both mom72 and mom19 can direct aac into mit ...19902177474
the general mitochondrial matrix processing protease from rat liver: structural characterization of the catalytic subunit.a critical step in the import of nuclear-encoded precursor proteins into mitochondria involves proteolytic cleavage of their amino-terminal leader peptides by processing proteases found in the mitochondrial matrix. we report here the characterization of the general matrix processing protease from rat liver mitochondria. the final enzyme preparation consisted of two polypeptides, a catalytically active 55-kda subunit and a 52-kda one. to deduce the complete primary structure of the 55-kda subunit ...19902236012
the development of a heterologous transformation system for the cellulolytic fungus trichoderma reesei based on a pyrg-negative mutant strain.six uridine auxotroph mutants of trichoderma reesei qm 9414 were isolated by resistance to 5-fluoroorotic acid and one strain was identified as omp-decarboxylase negative (pyr-) by a radiometric enzyme assay. transformation to uridine prototrophy was achieved with the pyr4 gene of neurospora crassa (up to 1500 transformants/micrograms) and with pyra of aspergillus niger (700-800 transformants/micrograms). in many transformants the pyr+ function seems to be present as extrachromosomal dna. there ...19902245476
mycenon, a new metabolite from a mycena species ta 87202 (basidiomycetes) as an inhibitor of isocitrate lyase.mycenon (c11h5cl3o3), a new inhibitor of isocitrate lyase (ec 4.1.3.1) was isolated from the culture broth of a basidiomycete, mycena sp. mycenon is a novel chlorinated benzoquinone derivative which is also active against bacteria and fungi. malate synthase (ec 4.1.3.2) the second key enzyme of the glyoxylate cycle was not affected by mycenon. isocitrate lyase preparations from plants, bacteria and fungi were sensitive. the following ki-values for mycenon have been determined: ricinus communis, ...19902258323
the purification and characterization of 3-dehydroquinase from streptomyces coelicolor.the enzyme 3-dehydroquinase was purified over 4000-fold to homogeneity from streptomyces coelicolor. the subunit mr estimated from polyacrylamide-gel electrophoresis in the presence of sds was 16,000. the native mr estimated by gel filtration on a superose 6 column was 209,000, indicating that the enzyme is a large oligomer. the enzyme was found to be extremely thermostable. this stability, along with the structural and kinetic properties of the enzyme, suggest that it is very similar to the qui ...19902306211
dna sequence analysis of the mitochondrial nd4l-nd5 gene complex from podospora anserina. duplication of the nd4l gene within its intron.a 15 kb region of the 100 kb mitochondrial genome of podospora anserina has been mapped and sequenced (1 kb = 10(3) base-pairs). the genes for nd4l and nd5 are identified as contiguous genes with overlapping termination and initiation codons. in race a (101 kb) the gene for nd4l (4.3 kb) has a gene duplication within an intron including a second subgroup ic intron. race s (95 kb) lacks this second gene complex. each intron has the identical 5' exon boundary. secondary structure analysis showed t ...19902319602
cloning and analysis of beta-tubulin gene from a protoctist.we have isolated and characterized by restriction endonuclease mapping, transcription pattern, and dna sequencing a beta-tubulin gene from the coenocytic freshwater protoctist, achlya klebsiana. the gene is intronless and has a single open reading frame that encodes a 444-amino acid residue polypeptide of mr 49,856. the protein shows a high degree of homology to other beta-tubulins, 85% identity to human beta-tubulin and 89% identity to beta-tubulin of the sporozoan (also a protoctist) plasmodiu ...19902394720
site-directed mutagenesis of the 'zinc finger' dna-binding domain of the nitrogen-regulatory protein nit2 of neurospora.the major nitrogen-regulatory gene nit-2 of neurospora crassa activates the expression of numerous unlinked structural genes which specify nitrogen-catabolic enzymes during conditions of nitrogen limitation. the nit-2 gene encodes a regulatory protein of 1036 amino acid residues with a single 'zinc finger' and a downstream basic region, which together may constitute a dna-binding domain. the zinc finger domain of the nit2 protein was synthesized in vitro and also expressed as a fusion protein in ...19902150539
neurospora endo-exonuclease is immunochemically related to the recc gene product of escherichia coli.immunochemical cross-reaction between the endo-exonuclease of neurospora crassa, an enzyme previously implicated in recombination and recombinational dna repair, and the recc-encoded polypeptide of escherichia coli has been detected by immunoblotting extracts of strains of e. coli having a deletion that includes the recbcd genes but carrying multicopy plasmids bearing all three of the recbcd genes or only one or two of these genes. it was predicted that homology would also be found at the amino ...19902152915
comparison and cross-species expression of the acetyl-coa synthetase genes of the ascomycete fungi, aspergillus nidulans and neurospora crassa.the genes encoding the acetate-inducible enzyme acetyl-coenzyme a synthetase from neurospora crassa and aspergillus nidulans (acu-5 and faca, respectively) have been cloned and their sequences compared. the predicted amino acid sequence of the aspergillus enzyme has 670 amino acid residues and that of the neurospora enzyme either 626 or 606 residues, depending upon which of the two possible initiation codons is used. the amino acid sequences following the second alternative aug show 86% homology ...19901972535
use of nuclear dna restriction fragment length polymorphisms to analyze the diversity of the aspergillus flavus group: a. flavus, a. parasiticus, and a. nomius.recombinant dna clones carrying high-copy or low-copy sequences from aspergillus nidulans and neurospora crassa were used to identify restriction fragment length polymorphisms (rflps) diagnostic for members of the a. flavus group: a. flavus, a. parasiticus, and a. nomius. these fungi were resolved into three distinct categories when they were grouped according to rflp patterns. subgroups within these categories were also evident. this limited rflp analysis of nuclear dna of members of the a. fla ...19901976300
molecular cloning of subunits of complex i from neurospora crassa. primary structure and in vitro expression of a 22-kda polypeptide.a lambda gt11 cdna expression library was screened with antibodies directed against individual subunits of complex i from neurospora crassa. clones encoding cytoplasmically synthesized polypeptides with apparent molecular masses of 22, 29, 31, and 33 kda were isolated. northern blot analysis revealed that the corresponding genes are transcribed into mrna species of about 0.85, 0.95, 1.3, and 1.4 kilobases, respectively. further characterization of clones encoding the 22-kda subunit was performed ...19902142943
tests for the genotoxicity of m-amsa, etoposide, teniposide and ellipticine in neurospora crassa.the antitumor agents m-amsa, etoposide, teniposide and ellipticine have been reported to be potent clastogens in mammalian cells but non- or weakly mutagenic in bacteria; these observations have been correlated to the interference of these chemicals with dna topoisomerase ii activity in the former, but not in the latter, organisms. the genotoxicity of these 4 agents was evaluated using ad-3 reverse- and forward-mutation tests in neurospora crassa. these agents (up to 0.8 mumole/plate) did not ca ...19902137196
remnants of an ancient pathway to l-phenylalanine and l-tyrosine in enteric bacteria: evolutionary implications and biotechnological impact.the pathway construction for biosynthesis of aromatic amino acids in escherichia coli is atypical of the phylogenetic subdivision of gram-negative bacteria to which it belongs (r. a. jensen, mol. biol. evol. 2:92-108, 1985). related organisms possess second pathways to phenylalanine and tyrosine which depend upon the expression of a monofunctional chorismate mutase (cm-f) and cyclohexadienyl dehydratase (cdt). some enteric bacteria, unlike e. coli, possess either cm-f or cdt. these essentially c ...19902082822
x-ray-induced specific-locus mutations in the ad-3 region of two-component heterokaryons of neurospora crassa. v. irreparable mutants of genotype ad-3a ad-3b, ad-3a ad-3b nic-2, and ad-3b nic-2 result from multilocus deletion and an unexpectedly high frequency of multiple-locus mutations.more extensive complementation tests than those performed initially (webber and de serres, 1965) on a series of 832 x-ray-induced specific-locus mutations in the adenine-3 (ad-3) region of a two-component heterokaryon (h-12) of neurospora crassa (de serres, 1989a) showed that unexpectedly high frequencies of specific-locus mutations in the ad-3 region have additional, but separate, sites of recessive lethal (rlcl) damage in the immediately adjacent genetic regions. the frequencies of these x-ray ...19902138248
molecular structure of a gene, vma1, encoding the catalytic subunit of h(+)-translocating adenosine triphosphatase from vacuolar membranes of saccharomyces cerevisiae.subunit a of the vacuolar membrane h(+)-translocating adenosine triphosphatase of the yeast saccharomyces cerevisiae contains a catalytic site for atp hydrolysis. n-terminal sequences of six tryptic peptides of the subunit were determined. based on the peptide sequence information, a 39-base oligonucleotide probe was synthesized, and the gene encoding the subunit (vma1) was isolated from a genomic dna library by hybridization. the nucleotide sequence of the gene predicts a polypeptide of 1,071 a ...19902139027
molecular analysis of nuc-1+, a gene controlling phosphorus acquisition in neurospora crassa.in response to phosphorus starvation, neurospora crassa makes several enzymes that are undetectable or barely detectable in phosphate-sufficient cultures. the nuc-1+ gene, whose product regulates the synthesis of these enzymes, was cloned and sequenced. the nuc-1+ gene encodes a protein of 824 amino acids with a predicted molecular weight of 87,429. the amino acid sequence shows homology with two yeast proteins whose functions are analogous to that of the nuc-1 protein. two nuc-1+ transcripts of ...19902146493
neurospora crassa a mating-type region.the mating-type locus of the haploid filamentous fungus neurospora crassa is a regulatory region that controls entry into the sexual cycle and prevents formation of mixed mating-type heterokaryons in the vegetative phase. the locus consists of alternative sequences called a and a. the a mating-type dna sequence of neurospora crassa is composed of a region of 5301 base pairs that has little similarity to the sequence present at the mating-type locus in an a mating-type strain. however, the sequen ...19902142303
deletion analysis of domain independence in the trp1 gene product of neurospora crassa.the trifunctional trp1 gene from neurospora crassa (n-trp1) was subcloned into the yeast-escherichia coli shuttle vector yep13 and expressed in saccharomyces cerevisiae. the three activities of the n-trp1 gene product were detected in yeast mutants that lacked either n-(5'-phosphoribosyl) anthranilate (pra) isomerase or both the glutamine amidotransferase function of anthranilate synthase and indole-3-glycerol phosphate (ingp) synthase. the protein was detected on immunoblots only as the full le ...19902147978
characterization of eukaryotic dna polymerases: aphidicolin resistant mutants of neurospora with altered dna polymerase.sucrose density gradient analysis of neurospora cell free extract showed at least three distinct peaks of enzyme activity; of these, a high molecular weight enzyme was identified as dna polymerase alpha because of its sensitivity to aphidicolin and to nem. dna polymerase mutants of neurospora crassa were isolated by their resistance to aphidicolin, a specific inhibitor of the eukaryotic dna polymerase alpha. some mutants showed an increase in the specific activity of the enzyme. one mutant (e-2- ...19902144049
neurospora crassa a mating-type region.the a mating-type region of neurospora crassa controls several major events in both the sexual and asexual phases of the fungal life cycle. this 3235-base-pair dna segment is not homologous to the comparable genetic region of the a mating type. the unique a and a regions are bordered by nearly identical dna sequences. the a genetic region contains at least two functional segments. one segment encodes a perithecium maturation function that is dependent on the second segment for phenotypic express ...19902142304
nit-2, the major positive-acting nitrogen regulatory gene of neurospora crassa, encodes a sequence-specific dna-binding protein.the nit-2 major nitrogen regulatory gene of neurospora crassa turns on the expression of various unlinked structural genes that specify nitrogen-catabolic enzymes under nitrogen-limitation conditions. the nit-2 gene encodes a protein of 1036 amino acid residues with a single zinc finger and a downstream basic region that may make up a dna-binding domain. the zinc-finger domain of the nit2 protein was synthesized in two ways to examine its dna-binding activity with gel-band-mobility shift and dna ...19902142530
primary structure of the non-transcribed spacer region and flanking sequences of the ribosomal dna of neurospora crassa and comparison with other organisms.the non-transcribed spacer (nts) region of the rdna of neurospora crassa contains the transcription regulatory sequences. we isolated a 3.4 kb ecori fragment from wild type n.crassa rdna and cloned in the plasmid pbr325 at the ecori site. the insert contains the entire nts region along with the flanking sequences. nucleotide sequencing of 3592 nt shows many interesting features like: the nts region is rich in g+c content (65% g+c); it contains the conserved rrna processing site 6 (with the nucle ...19902142594
paraquat toxicity and pyridine nucleotide coenzyme synthesis: a data correction.the decrease in pyridine nucleotide coenzymes which occurs during poisoning of escherichia coli by hyperbaric oxygen or paraquat is not due to impairment of nicotinatemononucleotide pyrophosphorylase (carboxylating) [ec 2.4.2.19] as was previously proposed (brown, o.r. et al. biochem. biophys. res. commun. 91:982-990; 1979). this was shown directly using extracts of e. coli, prepared after exposure to 1 mm paraquat or 4.2 atmospheres of oxygen. the enzyme also was not impaired in neurospora cras ...19902139629
structure, exon pattern, and chromosome mapping of the gene for cytosolic copper-zinc superoxide dismutase (sod-1) from neurospora crassa.a 4.8-kilobase bamhi-hindiii fragment encoding the entire neurospora crassa cuzn superoxide dismutase gene (herein designated sod-1) was isolated from a genomic library using two 60-base deoxyoligonucleotide probes corresponding to the published n. crassa amino acid sequence. the nucleotide sequence of the gene encodes an amino acid sequence matching the published protein sequence at 152 of 153 positions. codon preference shows an unusually strong bias such that only 32 of the possible 61 codons ...19902146266
heterozygous effects of x-ray-induced specific locus mutations in the ad-3 region of neurospora crassa: implications for human genetic risk assessment. 19902143295
carotenoid desaturases from rhodobacter capsulatus and neurospora crassa are structurally and functionally conserved and contain domains homologous to flavoprotein disulfide oxidoreductases.the characteristic red color of some photosynthetic bacteria and the orange color of neurospora conidia is due to the presence of carotenoids, photoprotective pigments synthesized by plants, algae, bacteria, and fungi. generally, carotenoids are tetraterpenes in which absorption of visible light and photoprotection are mediated by a chain of conjugated double bonds, the chromophore, which is formed by successive desaturations of phytoene, a colorless precursor. the genes al-1 and crti mediate th ...19902144293
two complex regions, including a tata sequence, are required for transcription by rna polymerase i in neurospora crassa.in order to define the rna polymerase i transcriptional apparatus and how it might interact with regulatory signals, the dna sequences necessary for 40s rrna transcription in neurospora crassa were determined. a systematic set of deletion, substitution and insertion mutations were assayed in a homologous in vitro system. the sequences required for transcription of the gene consist of two large domains (i and ii) from -113 to -37, and -29 to +4, respectively. complete deletion of either domain ab ...19902139932
molecular cloning and nucleotide sequence of full-length cdna for ascorbate oxidase from cultured pumpkin cells.a lambda gt11 cdna expression library was constructed from size-fractionated poly(a)-rich rna of cultured pumpkin cells. a full-length cdna clone for ascorbate oxidase mrna was selected from the library by screening with synthetic oligonucleotides designed from the amino-terminal sequence of ascorbate oxidase protein. the identity of the clone was confirmed by comparing the amino acid sequence deduced by nucleotide sequence analysis with that determined for the amino-terminal sequence of pumpkin ...19902143984
mating type and mating strategies in neurospora.in the heterothallic species neurospora crassa, strains of opposite mating type, a and a, must interact to give the series of events resulting in fruiting body formation, meiosis, and the generation of dormant ascospores. the mating type of a strain is specified by the dna sequence it carries in the mating type region; strains that are otherwise isogenic can mate and produce ascospores. the dna of the a and a regions have completely dissimilar sequences. probing dna from strains of each mating t ...19902140508
molybdenum cofactor requirement for in vitro activation of apo-molybdoenzymes of escherichia coli.the apo-nitrate reductase precursor in an escherichia coli chlb mutant preparation obtained following growth in the presence of tungstate is activated by incubation with protein fa and a heat-treated preparation from an e. coli crude extract. we show that the requirement for heat-treated e. coli crude extract can be fulfilled by material obtained from either of two heat-denatured purified e. coli molybdoenzymes, namely nitrate reductase or trimethylamine n-oxide reductase. apo-trimethylamine n-o ...19902141097
the neurospora crassa arg-2 locus. structure and expression of the gene encoding the small subunit of arginine-specific carbamoyl phosphate synthetase.we have characterized genomic and cdna clones for arg-2, the gene encoding the small subunit of the neurospora crassa arginine-specific carbamoyl phosphate synthetase (cps-a), and examined its transcriptional regulation. the polypeptide's predicted amino acid sequence (453 residues) is 56% and 36% identical with the sequences of the homologous polypeptides of saccharomyces cerevisiae and escherichia coli, respectively. the arg2 polypeptide has an additional amino-terminal domain with the hallmar ...19902141606
a neurospora crassa ribosomal protein gene, homologous to yeast cry1, contains sequences potentially coordinating its transcription with rrna genes.we have isolated and sequenced a neurospora crassa ribosomal protein gene (designated crp-2) strongly homologous to the rp59 gene (cry1) of yeast and the s14 ribosomal protein gene of mammals. the inferred sequence of the crp-2 protein is more homologous (83%) to the mammalian s14 sequence than to the yeast rp59 sequence (69%). the gene has three intervening sequences (ivss) two of which are offset 7 bp from the position of ivss in the mammalian genes. none correspond to the position of the ivs ...19901977135
neurospora crassa cdna clones coding for a new member of the ras protein family.a new member of the ras gene family was characterized from neurospora crassa cdna libraries. the clone designated nc-ras codes for a polypeptide containing 213 amino acids (mr 24,000). this polypeptide is 84% homologous to the h-ras-1 domain comprising the first 80 amino acids and 60% homologous to the next 84 residues. the nc-ras polypeptide contains all the well-known sequences involved in the interaction with gtp/gdp, the recognition of the y13-259 neutralizing antibody, the 'effector site' f ...19902146163
cys-3, the positive-acting sulfur regulatory gene of neurospora crassa, encodes a sequence-specific dna-binding protein.cys-3, the positive-acting master sulfur regulatory gene of neurospora crassa, turns on the expression of an entire set of unlinked structural genes which encode sulfur-catabolic enzymes. cys-3 encodes a protein of 236 amino acid residues and contains a potential bipartite dna-binding domain which consists of a leucine zipper and an adjacent highly basic region. gel band mobility shift and dna footprint experiments were used to demonstrate that the cys3 protein, expressed in escherichia coli, bi ...19902142156
evidence for the presence of a proton pump of the vacuolar h(+)-atpase type in the ruffled borders of osteoclasts.microsomal membrane vesicles prepared either from chicken medullary bone or isolated osteoclasts were shown to have atp-dependent h(+)-transport activity. this activity was n-ethylmaleimide-sensitive but resistant to oligomycin and orthovanadate, suggesting a vacuolar-type atpase. furthermore, immunological cross-reactivity of 60- and 70-kd osteoclast membrane antigens with neurospora crassa vacuolar atpase was observed when analyzed by immunoblotting. same antibodies labeled only osteoclasts in ...19902144003
expression in saccharomyces cerevisiae of a gene associated with cytoplasmic male sterility from maize: respiratory dysfunction and uncoupling of yeast mitochondria.we asked whether the mitochondrial t-urf13 gene, associated with the male sterility phenotype of t cytoplasm in maize, can be expressed in saccharomyces cerevisiae and whether this expression can mimic the effects observed in maize. we introduced the universal code equivalent of the t-urf13 gene into the s. cerevisiae nucleus by transformation and directed its translation product into mitochondria by means of a fusion with the targeting presequence from neurospora crassa at-pase subunit 9. we sh ...19902259341
an evolutionary comparison of acinetobacter calcoaceticus trpf with trpf genes of several organisms.the deduced amino acid sequence of acinetobacter calcoaceticus n-(5'-phosphoribosyl) anthranilate isomerase (prai), which is coded by trpf, was compared with trpf of caulobacter crescentus, escherichia coli, bacillus subtilis, saccharomyces cerevisiae, neurospora crassa, and aspergillus nidulans. sixty percent of identical or similar amino acids were located in alpha/beta tim (triose-phosphate isomerase) barrels and in residues important in substrate binding and catalysis. in addition, the analy ...19902299982
expression in yeast of the t-urf13 protein from texas male-sterile maize mitochondria confers sensitivity to methomyl and to texas-cytoplasm-specific fungal toxins.the mitochondrial gene t-urf13 from maize (zea mays l.) with texas male-sterile (t) cytoplasm codes for a unique 13 kd polypeptide, t-urf13, which is implicated in cytoplasmic male sterility and sensitivity to the insecticide methomyl and to host-specific fungal toxins produced by helminthosporium maydis race t (hmt toxin) and phyllosticta maydis (pm toxin). a chimeric gene coding for t-urf13 fused to the mitochondrial targeting peptide from the neurospora crassa atp synthase subunit 9 precursor ...19902303028
the 49 k subunit of nadh: ubiquinone reductase (complex i) from neurospora crassa mitochondria: primary structure of the gene and the protein.the primary structure of the 49 k subunit of the respiratory chain nadh:ubiquinone reductase (complex i) from neurospora crassa was determined by sequencing cdna, genomic dna and the n-terminus of the mature protein. the sequence lengths correlate to a molecular mass of 54,002 daltons for the preprotein and 49,239 daltons for the mature protein. the presequence consists of 42 amino acids of typical composition for sequences which target nuclear-encoded proteins into mitochondria. the mature prot ...19902147127
nucleotide sequence and analysis of nmr, a negative-acting regulatory gene in the nitrogen circuit of neurospora crassa.in neurospora the expression of a set of unlinked structural genes, which allows utilization of various nitrogen-containing compounds, is controlled by the positive-acting nit-2 gene and the negative-acting nmr gene. the nucleotide sequence of the nmr gene has been determined and a long open reading frame which encodes a putative protein of 54854 daltons has been identified. a full-length cdna clone was obtained and its the sequence revealed that the nmr gene contains no introns. the transcripti ...19902146484
the beta-tubulin gene of epichloë typhina from perennial ryegrass (lolium perenne).epichloë typhina is a biotrophic fungal pathogen which causes choke disease of pooid grasses. the anamorphic state, acremonium typhinum, is placed in the section albo-lanosa along with related, mutualistic, seed-disseminated endophytes. as an initial study of gene structure and evolution in epichloë and related endophytes, the beta-tubulin gene, tub2, of the perennial ryegrass choke pathogen (etprg) was cloned and sequenced. the coding sequence and the predicted beta-tubulin amino acid sequence ...19902147581
purification and further characterization of the second nitrate reductase of escherichia coli k12.two nitrate reductases, nitrate reductase a and nitrate reductase z, exist in escherichia coli. the nitrate reductase z enzyme has been purified from the membrane fraction of a strain which is deleted for the operon encoding the nitrate reductase a enzyme and which harbours a multicopy plasmid carrying the nitrate reductase z structural genes; it was purified 219 times with a yield of about 11%. it is an mr-230,000 complex containing 13 atoms iron and 12 atoms labile sulfur/molecule. the presenc ...19902139607
purification of chorismate synthase from a cell culture of the higher plant corydalis sempervirens pers.chorismate synthase (ec 4.6.1.4) was purified from a cell suspension culture of corydalis sempervirens almost 1000-fold to near homogeneity. the subunit mr estimated by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate was 41,900. the mr of the native enzyme was estimated to be 80,100 by gel filtration, suggesting a dimeric structure. antisera directed against the 41.9-kda protein also reacted with the native enzyme. further confirmation of the identity of the purified ...19902146922
induced expression of the aspergillus nidulans qute gene introduced by transformation into neurospora crassa.the qa-2 gene of neurospora crassa encodes catabolic dehydroquinase which catabolizes dehydroquinic acid to dehydroshikimic acid. the qute gene of aspergillus nidulans corresponds to the qa-2 gene of n. crassa. the plasmid peh1 containing the qute gene from a. nidulans was used to transform a qa-2- strain of n. crassa. in southern blot analyses, dnas isolated from these transformants hybridized specifically to the qute gene probe. northern blot analyses indicated that qute mrna was produced in t ...19902148798
expression of a neurospora crassa metallothionein and its variants in escherichia coli.the neurospora crassa metallothionein (nc) synthesis gene was cloned and expressed in escherichia coli in two different expression vectors (ping2 and pua7), both under the regulation of the salmonella typhimurium arabinose operon. upon induction with arabinose, the ping2-nc vector expressed as inclusion body-localized arab'::nc fusion protein of 21 kilodaltons. the pua7-nc vector expressed a 5.3-kilodalton lpp::nc fusion protein anchored to the outer membrane of the cell. cells expressing the nc ...19902148862
purification and characterization of catalase from a facultative alkalophilic bacillus.catalase was purified to an electrophoretically homogeneous state from the facultative alkalophilic bacterium, bacillus yn-2000, and some of its properties were studied. its molecular weight was 282,000 and its molecule was composed of four identical subunits. the enzyme contained two protoheme molecules per tetramer. the enzyme showed an absorption spectrum of typical high-spin ferric heme with a peak at 406 nm in the oxidized form and peaks at 440, 559, and 592 nm in the reduced form. in contr ...19902149854
atp-induced protyrosinase synthesis and carboxyl-terminal processing in neurospora crassa.the effects of 3'-5' cyclic amp and atp upon tyrosinase induction in neurospora crassa were examined. northern analysis of total cellular rna revealed rapid de novo synthesis of protyrosinase after addition of these substances to stationary-phase mycelia. the maturation of protyrosinase in crude extracts of mycelia was followed by western analysis. polyclonal rabbit antiserum directed against the denatured carboxyl-terminal extension of protyrosinase does recognize the proform and several interm ...19902150229
[genetic engineering in filamentous fungi: cloning of the invertase gene from neurospora crassa].the invertase wild type gene of n. crassa was cloned into the yrp7 yeast vector. this recombinant plasmid was selected by functional complementation of an invertaseless mutant strain of s. cerevisiae. the isolated recombinant plasmid (named pnc2) carries a 7.6 kb bamhi dna fragment from n. crassa. the cloned dna hybridized with the n. crassa genomic dna and transformed an invertase mutant of n. crassa inv- to inv+. transformation of n. crassa inv- to inv+ seems to take at least two different int ...19902151941
endo-exonuclease of aspergillus nidulans.endo-exonuclease (ee) has been found in both active and inactive, but trypsin-activatable, forms in aspergillus nidulans. active ee was present mainly in nuclei, mitochondria, and vacuoles, while trypsin-activatable ee was mainly in the cytosol. the active form accounts for over 90% of the neutral deoxyribonuclease activity extracted from mycelia. a single strand (ss) dna-binding ee associated with a 28 kilodalton (kda) polypeptide was partially purified and characterized. it was found to closel ...19902161674
spatial and biological characterisation of the complete quinic acid utilisation gene cluster in aspergillus nidulans.heterologous probing of restriction digests of chromosomal dna from aspergillus nidulans with radioactively labelled probes encoding dehydroshikimate dehydratase (qa-4) and a repressor gene (qa1-s) from neurospora crassa revealed a pattern of hybridisation inconsistent with an equivalent single copy of each gene in a. nidulans. screening of size-selected and total genome a. nidulans dna libraries allowed the isolation of four unique classes of sequence, two of which hybridised to the qa-4 probe, ...19902175387
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