SEARCH · Search PubMed

Results for “Niger”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

SourceResearch connectionFrom yearThrough year

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 127 records · Page 7Linked to original sources

PubMed1997

Stability and identification of active-site residues of carboxymethylcellulases from Aspergillus niger and Cellulomonas biazotea.

Determination of the apparent pKa's of purified carboxymethylcellulases from Aspergillus niger and Cellulomonas biazotea at different temperatures and in the presence of dioxane indicated two side chain carboxyl groups which controlled the limiting rate in both organisms. The thermostability of both enzymes slightly decreased with increasing pH from 5 to 75 but was unaffected in the presence of 0.5 mmol/L Mn2+. The CMCase from C. biazotea had an activation energy of 35 kJ/mol and a half-life of 89 min in the presence of 8 mol/L urea at 40 degrees C. The half-life of CMCase from A. niger in 8 mol/L urea and at 37 degrees C was 125 min as determined by a 0-9 mol/L transverse urea gradient PAGE. The CMCases from A. niger and C. biazotea had the same thermostabilities in the absence of CMC although the enzyme from the former was more thermostable in the presence of the substrate. The CMCase from A. niger was also more efficient in hydrolyzing CMC than the enzyme from C. biazotea.

Actinomycetales↗

PubMed1997

Production of the homologous pectin lyase B protein in six genetically defined protease-deficient Aspergillus niger mutant strains.

An expression cassette has been transformed into six protease-deficient (prt) mutant strains of Aspergillus niger. Transformants were tested for improved production of the proteolytically susceptible PELB tester protein. In four complementation groups (prtA, B, D and F) distinct improvement of PELB yield was observed. These in vivo experiments in single prt mutants confirmed earlier in vitro PELB degradation data and demonstrated how the use of protease-deficient mutants can significantly improve protein production in A. niger. The strong effects of several prt alleles on the stability of the PELB tester protein have initiated a more detailed genetical and molecular characterization of the prt mutations. Mapping of the cloned protease genes pepA [I], B [II], C [IV], D [I], E [IV] and F [IV] indicated that none of the prt mutations represent alleles of the presently cloned protease (pep) genes from A. niger. Analysis of the expression of the pep genes in prt strains demonstrated that the strongly reduced protease activities observed in several prt mutants are not reflected by reduced transcription levels for a number of extracellular proteases. These results indicate that the mode of action of the prt genes constitute an interesting group of new genetic functions which severely affect protease production, and as such improve protein production, in A. niger.

Aspergillus niger↗

PubMed1998

Genetic analysis of resistance to fenpropimorph in Aspergillus niger.

Resistance to the morpholine-fungicide fenpropimorph was studied in Aspergillus niger and A. nidulans. Mass selection of conidia of A. nidulans on agar amended with the fungicide at different concentrations did not yield of resistant mutants, even after UV-treatment of the conidia. In contrast, similar experiments with A. niger generated many fenpropimorph-resistant mutants. The mutants displayed cross-resistance to fenpropidin and generally showed wild-type sensitivity to the unrelated toxicants fenarimol and cycloheximide. Genetic analysis of fenpropimorph resistance in A. niger was carried out by means of the parasexual cycle. In the mutants tested, two genes located on linkage group II were involved in fenpropimorph resistance. Dominance tests showed that resistance to fenpropimorph in A. niger is recessive.

Aspergillus niger↗

PubMed1996

The transposable element Tan1 of Aspergillus niger var. awamori, a new member of the Fot1 family.

Aspergillus niger var. awamori has transposable elements that we refer to as Vader and Tan1 (transposon A. niger). Vader was identified by screening unstable nitrate reductase (niaD) mutants for insertions. Four of the isolated niaD mutants were shown to contain a small insertion element. This 437 bp insertion element, Vader, is flanked by 44 bp inverted repeats (IR) and is present in approximately 15 copies in the genomes of two A. niger strains examined. A synthetic 44 bp oligomer of the inverted repeat of Vader has now been used to clone, via the polymerase chain reaction, a 2.3 kb Tan1 element. The Tan1 element has also been isolated from a partial genomic library. Tan1 is present as a single copy in A. niger var. awamori. The Tan1 element has a unique organization: IR-ORF-IR-IR-Vader-IR. The single open reading frame (ORF) (1668 bp) encodes a putative transposase homologous to Fusarium oxysporum Fot1 and Magnaporthe grisea Pot2. Immediately 3' to the second inverted repeat, which bounds the transposase, is a copy of the AT-rich Vader element. We hypothesize that at some stage the independent Vader element, although inactive by itself, arose from Tan1, resulting in current strains with only one copy of Tan1 providing transposase activity and numerous mobile copies of Vader dispersed in the genome.

Amino Acid Sequence↗

PubMed2001

Isolation of UV-induced mutations in the areA nitrogen regulatory gene of Aspergillus niger, and construction of a disruption mutant.

The areA gene of filamentous fungi encodes a positive-acting transcriptional factor required for the expression of genes involved in the utilisation of nitrogen sources other than ammonium and glutamine. In Aspergillus niger we have isolated three UV-induced areA mutants and constructed a well-defined disruption allele of the areA gene. The areA gene was genetically localised on Linkage Group III, 3.6 map units (m.u.) from bioA1 and 4.5 m.u. from lysA7. Analysis of the expression of the nitrate reductase encoding gene and of nitrate reductase activities show that the mutated areA strains behave as loss-of-function mutants and can be classified as areAr type. In addition, growth tests were performed using several nitrogen sources in combination with glucose. The results suggest that, unlike the case in A. nidulans, in A. niger the AreA protein also plays a role in the presence of ammonium. Furthermore, the spectrum of protease activities secreted by A. niger differs from that produced by A. nidulans, as only A. niger is able to degrade elastin.

Aspergillus niger↗

PubMed2001

The Aspergillus niger cypA gene encodes a cyclophilin that mediates sensitivity to the immunosuppressant cyclosporin A.

Here we report the cloning and characterization of a gene, cypA, from Aspergillus niger that encodes a peptidyl prolyl cis-trans isomerase (PPIase) belonging to the cyclophilin family. Sequencing of both genomic and cDNA clones revealed two ORFs in cypA, one encoding a 19-kDa protein of 174 amino acid residues and the other a 24-kDa protein of 219 amino acid residues, with overall identities of 27-77% to the homologous cyclophilins from prokaryotic and eukaryotic organisms. Expression of the 19-kDa CYPA-(His)(6) in E. coli shows that the purified protein has PPIase activity which is inhibited by cyclosporin A. Northern analysis shows two specific cypA transcripts, the smaller transcript encodes the cytosolic 19-kDa CYPA protein, the larger transcript encodes the putative mitochondrial 24 kDa CYPA protein. The transcript for the cytosolic CYPA is expressed at a higher basal level than that for the mitochondrial protein. The presence of tunicamycin, DTT or cyclosporin A in the medium does not affect the expression level of cypA. Its expression is however slightly induced by heat shock. Growing A. niger mycelium in the presence of cyclosporin A leads to an increase in hyphal branching prior to growth arrest. Overexpression of cypA under the control of its own promoter in A. niger results in increased sensitivity to cyclosporin A, suggesting that cypA encodes the cellular target for cyclosporin A in A. niger.

Amino Acid Sequence↗

PubMedDate not supplied

Isolation and sequencing of a new glucoamylase gene from an Aspergillus niger aggregate strain (DSM 823) molecularly classified as Aspergillus tubingensis.

Based on morphological characteristics the taxa included in the Aspergillus aggregate can hardly be differentiated. For that reason the phylogeny of this genus was revised several times as different criteria, from morphological to later molecular, were used. We found, comparing nucleotide sequences of the ITS-region, that the strain Aspergillus niger (DSM 823) which is claimed to be identical to the strains ATCC 10577, IMI 027809, NCTC 7193 and NRRL 2322 can be molecularly classified as Aspergillus tubingensis, exhibiting 100% identity with the A. tubingensis CBS strains 643.92 and 127.49. We amplified, cloned and sequenced a new glucoamylase gene (glaA) from this strain of A. tubingensis (A. niger DSM 823) using primers derived from A. niger glucoamylase G1. The amplified cDNA fragment of 2013 bp contained an open reading frame encoding 648 amino acid residues. The calculated molecular mass of the glucoamylase, deduced from the amino acid sequence, was 68 kDa. The nucleotide sequence of glaA showed 99% similarity with glucoamylases from Aspergillus kawachii and Aspergillus shirousami, whereas the similarity with the glucoamylase G1 from A. niger was 92%

Amino Acid Sequence↗

PubMed1993

Secretion of heterologous proteins by Aspergillus niger: production of active human interleukin-6 in a protease-deficient mutant by KEX2-like processing of a glucoamylase-hIL6 fusion protein.

To develop improved methods for heterologous protein production in Aspergillus niger, we studied the secretion of human interleukin-6 (hIL6). Since in vitro experiments with culture medium revealed that hIL6 was rapidly degraded, several protease-deficient strains of A. niger were isolated and tested for reduced degradation of hIL6 compared with the wild-type strain. The mutant strain giving the least degradative effect on hIL6 (designated AB1.13) was transformed with several hIL6-expression plasmids. Initially, hIL6 was expressed using various signal sequences fused to the sequence of mature hIL6. The resulting transformants did not produce detectable amounts of hIL6, despite high transcription levels in one transformant. We hypothesized that hIL6 was not efficiently processed during passage along the secretion pathway. Therefore, hIL6 was expressed as a fusion protein with glucoamylase, a protein which is efficiently secreted by A. niger and expression of which can easily be measured enzymatically. To obtain mature hIL6, a sequence encoding the KEX2 cleavage-site (Lys-Arg) was inserted between glucoamylase and hIL6 sequences. Mature active hIL6 was found to be secreted in the extracellular medium. Using this combined approach of transforming a protease-deficient strain with a fusion construct containing the KEX2 site, up to 15 mg l-1 active hIL6 was obtained in shake-flask culture. A fusion construct without the KEX2 site resulted in substantially higher production of the fusion protein, but hIL6 was not active in the fused form. These results indicate that A. niger contains a protease with similar specificity as the KEX2 protease from yeast.

Amino Acid Sequence↗

PubMed1985

Isolation and characterization of the Aspergillus niger trpC gene.

The Aspergillus niger trpC gene was isolated by complementation experiments with an Escherichia coli trpC mutant. Plasmid DNA containing the A. niger trpC gene transforms an Aspergillus nidulans mutant strain, defective in all three enzymatic activities of the trpC gene, to Trp+, indicating the presence of a complete and functional trpC gene. Southern blot analysis of DNA from these Trp+ transformants showed that plasmid DNA was present but that this DNA was not integrated at the site of the chromosomal trpC locus. The A. niger trpC gene was localized on the cloned fragment by heterologous hybridization experiments and sequence analysis. These experiments suggest that the organization of the A. niger trpC gene is identical to that of the analogous A. nidulans trpC and the Neurospora crassa trp-1 genes.

Aldose-Ketose Isomerases↗

PubMed1989

Transformation of Aspergillus niger with the homologous nitrate reductase gene.

A homologous transformation for Aspergillus niger was developed based on the nitrate reductase structural gene niaD. This system offered certain advantages over existing A. niger systems, such as the ease of recipient mutant isolation, absence of abortive transformants, convenient enzyme assay, ease of transformant stability testing, and complete absence of background growth. Transformation frequencies of up to 100 transformants per microgram DNA were obtained with the vector pSTA10 which carries the niaD gene of A. niger. Southern blotting analysis indicated that vector DNA had integrated into the genome of A. niger. Mitotic stability studies demonstrated that while some transformants were as stable as the wild-type (wt), others were markedly less so. No correlation was seen between plasmid integration, mitotic stability and nitrate reductase activity, which was markedly different from wt in only three of the transformants examined.

Aspergillus niger↗

PubMed2005

Identification of an anonymous RFLP DNA probe through multiple arbitrary amplicon profiling and its use for strain differentiation in a field isolate of cellulose-degrading Aspergillus niger.

The genome of Aspergillus niger (MPS-002) was subjected to RAPD fingerprinting using none different random oligonucleotide primers. A 0.7 Kb PCR amplicon, generated by primer-3 could be used as a RFLP probe to differentiate A. niger (ATCC 16880) from A. niger (MPS-002). The probe revealed DNA polymorphism internal to two different EcoRI recognition sequences spaced apart at a distance of 0.4 Kb within a 4.0 Kb EcoRI fragment of the genome of both the strains. Localized genome mapping analysis further revealed that the 0.7 Kb RFLP probe was positioned at a distance of 2.7 Kb and 0.6 Kb from the two ends of a 4.0 Kb EcoRI fragment, respectively within the genome of the two strains of A. niger.

Aspergillus niger↗

PubMed2006

RFLP characterization of Aspergillus niger aggregate species from grapes from Europe and Israel.

In order to characterize by molecular methods the Aspergillus niger aggregate species involved in the ochratoxin A (OTA) contamination of European wine grapes and table grapes from Israel, a total of 173 strains were studied. The ITS-5.8S rDNA fragments of 173 A. niger agreggate strains from grapes included in this study were amplified and their PCR amplicons were RsaI digested in order to classify the strains in the RFLP types, N and T. All of the strains belonging to the A. niger aggregate were classified into the two RFLP types previously defined: type N (43%) and type T (57%). Twenty out of the 173 strains of A. niger aggregate produced OTA (0.1 to 10.5 mug g(-1)). All the OTA producing species belonged to the N-RFLP type.

Aspergillus niger↗

PubMed2004

Homologous expression of the feruloyl esterase B gene from Aspergillus niger and characterization of the recombinant enzyme.

The faeB gene encoding the feruloyl esterase B (FAEB) was isolated from Aspergillus niger BRFM131 genomic DNA. The faeB gene, with additional sequence coding for a C-terminal histidine tag, was inserted into an expression vector under the control of the gpd promoter and trpC terminator and expressed in a protease deficient A. niger strain. Homologous overproduction allows to reach an esterase activity of 18 nkat mL(-1) against MCA as substrate. The improvement factor was 16-fold higher as compared to the production level obtained with non-transformed A. niger strain induced by sugar beet pulp. The corresponding secretion yield was estimated to be around 100 mg L(-1). Recombinant FAEB was purified 14.6-fold to homogeneity from an 8-day-old culture by a single affinity chromatographic step with a recovery of 64%. SDS-PAGE revealed a single band with a molecular mass of 75 kDa, while under non-denatured conditions, native enzyme has a molecular mass of around 150 kDa confirming that the recombinant FAEB is a homodimer. The recombinant and native FAEB have the same characteristics concerning temperature and pH optima, i.e., 50 degrees C and 6, respectively. In addition, the recombinant FAEB was determined to be quite stable up to 50 degrees C for 120 min. Kinetic constants for MCA, MpCA, and chlorogenic acid (5-O-caffeoyl quinic acid) were as follows: Km: 0.13, 0.029, and 0.16 mM and Vmax: 1101, 527.6, and 28.3 nkat mg(-1), respectively. This is the first report on the homologous overproduction of feruloyl esterase B in A. niger.

Aspergillus niger↗

PubMed1998

Aberrant processing of wild-type and mutant bovine pancreatic trypsin inhibitor secreted by Aspergillus niger.

Bovine pancreatic trypsin inhibitor (BPTI) was secreted by Aspergillus niger at yields of up to 23 mg l-1 using a protein fusion strategy. BPTI was linked to part of the fungal glucoamylase protein (GAM) with a dibasic amino acid (KEX2) processing site at the fusion junction. Electrospray ionisation mass spectrometry and N-terminal protein sequencing revealed that, although biologically active in vitro, the purified products from a number of independent transformants consisted of a mixture of BPTI molecules differing at the N-terminus. Approximately 35-60% of this mixture was processed correctly. Aberrant processing of the GAM-BPTI fusion protein by the A. niger KEX2-like endoprotease was the most likely cause of this variation although the involvement of other fungal endoproteases could not be ruled out. In vitro studies have highlighted a weak interaction between BPTI and the Saccharomyces cerevisiae KEX2 endoprotease, suggesting that BPTI is not a potent inhibitor of KEX2p. A small proportion of the recombinant BPTI (10%) showed 'nicking' of the K15-A16 bond, indicating an interaction with a fungal trypsin-like enzyme. Mutant BPTI homologues designed to have anti-elastase activity, BPTI(K15V), BPTI(K15V,P13I) and BPTI(K15V,G12A), have also been expressed and secreted by A. niger. They also showed a similar spectrum of aberrant N-terminal processing but no 'nicking' of the K15-V16 bond was observed. Comparison of A. niger with other expression systems showed that it is an effective system for producing BPTI and its homologues, although not all molecules were correctly processed. This variation in processing efficiency may be useful in understanding the important determinants of protein processing in this fungus.

Amino Acid Sequence↗

PubMed1999

Biotransformation of (1-phenyl)ethyl hydroperoxide with Aspergillus niger: a model study on enzyme selectivity and on the induction of peroxidase activity.

The biocatalytic enantioselective reduction of (1-phenyl)ethyl hydroperoxide (1) by the fungus Aspergillus niger to the corresponding alcohol 2 involves a multi-enzyme biotransformation of the hydroperoxide 1, as revealed by the change in the enantioselectivity as a function of incubation times. This unusual behavior is not exhibited by other fungi and seems to be restricted to A. niger. Furthermore, the peroxidase and other oxidoreductase activities of A. niger depend on the availability of metal ions such as Fe2+, Mn2+ and Zn2+ in the growth medium, since the addition of Fe2+ ions substantially (threefold) increases the enantioselectivity, whereas addition of Mn2+ and Zn2+ ions decreases it. Finally, the cold shock (4 degrees C) significantly enhances the reduction of the hydroperoxide by the microorganism A. niger.

Aspergillus niger↗

PubMed1999

Glucoamylase overexpression and secretion in Aspergillus niger: analysis of glycosylation.

We have studied the effects of overexpression and secretion of a homologous model glycoprotein, glucoamylase (GAM-1), on glycosylation in a single gene copy wild-type parent and multiple gene copy transformants of Aspergillus niger. In batch culture the B36 strain, which possess 80 additional copies of the GAM glaA gene, secreted about 5-8-fold more protein and GAM-1 than the parent strain (N402). A comparison of the glycosylation of GAM-1 secreted by the parent strain with that secreted by the multiple copy and hyper-secreting B36 strain showed that both the N-linked and O-linked glycan composition was very similar. Short oligomannose N-linked glycans were found (Man(7-8)GlcNAc(2)). O-Linked glycans were comprised of short (1-3 residues) oligosaccharide chains of mannose and galactose. Evidence is presented that this galactose is present in the novel galactofuranose conformation. This glycan composition of GAM-1 differed from that of a commercially available (A. niger) GAM source. Microsomes prepared from the mycelium showed a 2-3-fold co-ordinated increase in the activity of the dolichol phosphate:glycosyltransferases. Similar results were obtained from strains B1 (20 copies of glaA) and N402 when grown at a low dilution rate in a chemostat, although both the levels of GAM secretion and the activities of the dolichol phosphate:glycosyltransferases were lower than found in batch culture. These data suggest that A. niger is capable of secreting large amounts of a single glycoprotein combined with higher activity levels of the dolichol phosphate:glycosyltransferases without an increase in the heterogeneity of the glycan structures. Thus, from a biotechnological viewpoint, protein glycosylation may not be a bottleneck to enhanced glycoprotein production using A. niger.

Aspergillus niger↗

PubMed2001

Aspergillus niger absorbs copper and zinc from swine wastewater.

Wastewater from swine confined-housing operations contains elevated levels of copper and zinc due to their abundance in feed. These metals may accumulate to phytotoxic levels in some agricultural soils of North Carolina due to land application of treated swine effluent. We evaluated fungi for their ability to remove these metals from wastewater and found Aspergillus niger best suited for this purpose. A. niger was able to grow on plates amended with copper at a level five times that inhibitory to the growth of Saccharomyes cerevisiae. We also found evidence for internal absorption as the mechanism used by A. niger to detoxify its environment of copper, a property of the fungus that has not been previously exploited for metal bioremediation. In this report, we show that A. niger is capable of removing 91% of the copper and 70% of the zinc from treated swine effluent.

Animals↗

PubMed2004

Production of cellulases and hemicellulases by Aspergillus niger KK2 from lignocellulosic biomass.

To investigate the production of cellulases and hemicellulases from Aspergillus niger KK2, solid state fermentation (SSF) was performed by using different ratios of rice straw and wheat bran. When A. niger KK2 was grown on rice straw alone as a solid support in SSF, the maximum FPase activity was 19.5 IU g(-1) in 4 days. Also, CMCase (129 IU g(-1)), beta-glucosidase (100 IU g(-1)), xylanase (5070 IU g(-1)) and beta-xylosidase (193 IU g(-1)) activities were concurrently obtained after 5-6 days of fermentation. The higher enzyme activities produced by A. niger KK2 is a significant advantage from the viewpoint of practical saccharification reaction. Cellulases and hemicellulases produced by A. niger KK2 might be applied to pulp and paper industry, feed industry and chemical industry.

Aspergillus niger↗