Aspergillus oryzae strains with a large deletion of the aflatoxin biosynthetic homologous gene cluster differentiated by chromosomal breakage

ABSTRACT We analyzed the LKP fimbrial gene clusters of six piliated strains of a cryptic genospecies of Haemophilus isolated from the genital tracts of adult patients (five strains) and from an infected neonate. In a group of 19 genital strains, LKP-like genes have been found in only these 6 strains. In addition to the ghfA, ghfD, and ghfE genes previously described, we characterized two genes, designated ghfB and ghfC, encoding the putative chaperone and assembly platform proteins. All six strains had a complete and unique LKP-like gene cluster consisting of the five genes ghfA to ghfE, homologous to genes hifA to hifE of Haemophilus influenzae. The sequences of the coding and intergenic regions of the ghf clusters of the six strains were remarkably homologous. Unlike hif clusters, which are inserted between purE and pepN, the ghf cluster was inserted between purK and pepN on the chromosome. Analysis of the flanking regions of the ghf cluster identified a large deletion, identical in the 5′ end regions of all strains, including the whole purE gene and much of the purK gene. Ultrastructural observations, an attempt at enriching LKP fimbriae, and hemagglutination experiments demonstrated that none of the strains had LKP-type fimbriae. Nevertheless, reverse transcription (RT)-PCR showed that ghf genes were transcribed in four of the six strains. Sequencing of the intergenic ghfA-ghfB regions, including the ghf gene promoters, showed that the absence of transcripts in the remaining two strains was due to a decrease in the number of TA repeats (4 or 9 repeats rather than 10) between the −10 and −35 boxes of the two overlapping and divergent promoters. The other four strains, which had ghf transcripts, had the optimal 10 TA repeats (one strain) or 5 repeats associated with putative alternative −35 boxes (three strains). The absence of 10 repeated palindromic sequences of 44 or 45 nucleotides upstream of ghfB induces an increased instability of mRNA, as quantified by real-time RT-PCR, and may explain why the LKP fimbrial gene cluster is not expressed in these strains.

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Effects of Chloromethanes on Growth of and Deletion of the pce Gene Cluster in Dehalorespiring Desulfitobacterium hafniense Strain Y51

Applied and Environmental Microbiology ◽

10.1128/aem.00979-06 ◽

2006 ◽

Vol 72 (9) ◽

pp. 5998-6003 ◽

Cited By ~ 29

Author(s):

Taiki Futagami ◽

Takehito Yamaguchi ◽

Shun-ichi Nakayama ◽

Masatoshi Goto ◽

Kensuke Furukawa

Keyword(s):

Carbon Tetrachloride ◽

Growth Inhibition ◽

Gene Cluster ◽

Inhibitory Effect ◽

Large Deletion ◽

Growth Inhibitory Effect ◽

Reductive Dehalogenase ◽

Growth Inhibitory ◽

Desulfitobacterium Hafniense ◽

Significant Growth Inhibition

ABSTRACT The dehalorespiring Desulfitobacterium hafniense strain Y51 efficiently dechlorinates tetrachloroethene (PCE) to cis-1,2-dichloroethene (cis-DCE) via trichloroethene by PceA reductive dehalogenase encoded by the pceA gene. In a previous study, we found that the significant growth inhibition of strain Y51 occurred in the presence of commercial cis-DCE. In this study, it turned out that the growth inhibition was caused by chloroform (CF) contamination of cis-DCE. Interestingly, CF did not affect the growth of PCE-nondechlorinating SD (small deletion) and LD (large deletion) variants, where the former fails to transcribe the pceABC genes caused by a deletion of the promoter and the latter lost the entire pceABCT gene cluster. Therefore, PCE-nondechlorinating variants, mostly LD variant, became predominant, and dechlorination activity was significantly reduced in the presence of CF. Moreover, such a growth inhibitory effect was also observed in the presence of carbon tetrachloride at 1 μM, but not carbon dichloride even at 1 mM.

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Heterologous Biosynthesis and Genomics-Driven Derivatization of Fungal Bioactive Sesterterpenoid Variecolin

10.33774/chemrxiv-2021-x171c ◽

2021 ◽

Author(s):

Dexiu Yan ◽

Jemma Arakelyan ◽

Teng Wan ◽

Tsz Ki Chan ◽

Dohyun Ahn ◽

...

Keyword(s):

Cytochrome P450 ◽

Anticancer Activity ◽

Gene Cluster ◽

Aspergillus Oryzae ◽

Biosynthetic Gene Cluster ◽

Heterologous Production ◽

Biosynthetic Gene ◽

Aspergillus Aculeatus ◽

Heterologous Biosynthesis

The biosynthetic gene cluster of fungal bioactive sesterterpenoids, variecolin (1) and variecolactone (2), was identified in Aspergillus aculeatus ATCC 16872. Heterologous production of 1 and 2 was achieved in Aspergillus oryzae by expressing the sesterterpene synthase VrcA and the cytochrome P450 VrcB. Intriguingly, the replacement of VrcB with homologous P450s from other fungal terpenoid pathways yielded three new variecolin analogues, one of which exhibited potent anticancer activity comparable to that of 1.

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Efficient Reconstitution of Basidiomycota Diterpene Erinacine Gene Cluster in Ascomycota Host Aspergillus oryzae Based on Genomic DNA Sequences

Journal of the American Chemical Society ◽

10.1021/jacs.9b08935 ◽

2019 ◽

Vol 141 (39) ◽

pp. 15519-15523 ◽

Cited By ~ 6

Author(s):

Chengwei Liu ◽

Atsushi Minami ◽

Taro Ozaki ◽

Jing Wu ◽

Hirokazu Kawagishi ◽

...

Keyword(s):

Gene Cluster ◽

Aspergillus Oryzae ◽

Dna Sequences ◽

Genomic Dna

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Identification of the Monooxygenase Gene Clusters Responsible for the Regioselective Oxidation of Phenol to Hydroquinone in Mycobacteria

Applied and Environmental Microbiology ◽

10.1128/aem.02316-10 ◽

2010 ◽

Vol 77 (4) ◽

pp. 1214-1220 ◽

Cited By ~ 21

Author(s):

Toshiki Furuya ◽

Satomi Hirose ◽

Hisashi Osanai ◽

Hisashi Semba ◽

Kuniki Kino

Keyword(s):

Gene Cluster ◽

Large Subunit ◽

Gene Clusters ◽

Small Subunit ◽

Homologous Gene ◽

Oxidation Activity ◽

Monooxygenase Gene ◽

Regioselective Oxidation ◽

Binuclear Iron ◽

Induced Proteins

ABSTRACTMycobacterium goodiistrain 12523 is an actinomycete that is able to oxidize phenol regioselectively at theparaposition to produce hydroquinone. In this study, we investigated the genes responsible for this unique regioselective oxidation. On the basis of the fact that the oxidation activity ofM. goodiistrain 12523 toward phenol is induced in the presence of acetone, we first identified acetone-induced proteins in this microorganism by two-dimensional electrophoretic analysis. The N-terminal amino acid sequence of one of these acetone-induced proteins shares 100% identity with that of the protein encoded by the open reading frame Msmeg_1971 inMycobacterium smegmatisstrain mc2155, whose genome sequence has been determined. Since Msmeg_1971, Msmeg_1972, Msmeg_1973, and Msmeg_1974 constitute a putative binuclear iron monooxygenase gene cluster, we cloned this gene cluster ofM. smegmatisstrain mc2155 and its homologous gene cluster found inM. goodiistrain 12523. Sequence analysis of these binuclear iron monooxygenase gene clusters revealed the presence of four genes designatedmimABCD, which encode an oxygenase large subunit, a reductase, an oxygenase small subunit, and a coupling protein, respectively. When themimAgene (Msmeg_1971) ofM. smegmatisstrain mc2155, which was also found to be able to oxidize phenol to hydroquinone, was deleted, this mutant lost the oxidation ability. This ability was restored by introduction of themimAgene ofM. smegmatisstrain mc2155 or ofM. goodiistrain 12523 into this mutant. Interestingly, we found that these gene clusters also play essential roles in propane and acetone metabolism in these mycobacteria.

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Operon for Biosynthesis of Lipstatin, the Beta-Lactone Inhibitor of Human Pancreatic Lipase

Applied and Environmental Microbiology ◽

10.1128/aem.01765-14 ◽

2014 ◽

Vol 80 (24) ◽

pp. 7473-7483 ◽

Cited By ~ 18

Author(s):

Tingli Bai ◽

Daozhong Zhang ◽

Shuangjun Lin ◽

Qingshan Long ◽

Yemin Wang ◽

...

Keyword(s):

Fatty Acid ◽

Pancreatic Lipase ◽

Acyl Carrier Protein ◽

Gene Knockout ◽

Single Gene ◽

Oxidative Degradation ◽

Large Deletion ◽

Homologous Gene ◽

Content Type ◽

Dihydroxy Fatty Acid

ABSTRACTLipstatin, isolated fromStreptomyces toxytricinias a potent and selective inhibitor of human pancreatic lipase, is a precursor for tetrahydrolipstatin (also known as orlistat, Xenical, and Alli), the only FDA-approved antiobesity medication for long-term use. Lipstatin features a 2-hexyl-3,5-dihydroxy-7,10-hexadecadienoic-β-lactone structure with anN-formyl-l-leucine group attached as an ester to the 5-hydroxy group. It has been suggested that the α-branched 3,5-dihydroxy fatty acid β-lactone moiety of lipstatin inS. toxytriciniis derived from Claisen condensation between two fatty acid substrates, which are derived from incomplete oxidative degradation of linoleic acid based on feeding experiments. In this study, we identified a six-gene operon (lst) that was essential for the biosynthesis of lipstatin by large-deletion, complementation, and single-gene knockout experiments.lstA,lstB, andlstC, which encode two β-ketoacyl–acyl carrier protein synthase III homologues and an acyl coenzyme A (acyl-CoA) synthetase homologue, were indicated to be responsible for the generation of the α-branched 3,5-dihydroxy fatty acid backbone. Subsequently, the nonribosomal peptide synthetase (NRPS) genelstEand the putative formyltransferase genelstFwere involved in decoration of the α-branched 3,5-dihydroxy fatty acid chain with an N-formylated leucine residue. Finally, the 3β-hydroxysteroid dehydrogenase-homologous genelstDmight be responsible for the reduction of the β-keto group of the biosynthetic intermediate, thereby facilitating the formation of the unique β-lactone ring.

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Two Different Lantibiotic-Like Peptides Originate from the Ericin Gene Cluster of Bacillus subtilis A1/3

Journal of Bacteriology ◽

10.1128/jb.184.6.1703-1711.2002 ◽

2002 ◽

Vol 184 (6) ◽

pp. 1703-1711 ◽

Cited By ~ 90

Author(s):

Torsten Stein ◽

Stefan Borchert ◽

Birgit Conrad ◽

Jörg Feesche ◽

Brigitte Hofemeister ◽

...

Keyword(s):

Bacillus Subtilis ◽

Gene Cluster ◽

High Performance ◽

Genetic Locus ◽

Reversed Phase ◽

Homologous Gene ◽

Dependent Manner ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Tof Ms

ABSTRACT A lantibiotic gene cluster was identified in Bacillus subtilis A1/3 showing a high degree of homology to the subtilin gene cluster and occupying the same genetic locus as the spa genes in B. subtilis ATCC 6633. The gene cluster exhibits diversity with respect to duplication of two subtilin-like genes which are separated by a sequence similar to a portion of a lanC gene. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) analyses of B. subtilis A1/3 culture extracts confirmed the presence of two lantibiotic-like peptides, ericin S (3,442 Da) and ericin A (2,986 Da). Disruption of the lanB-homologous gene eriB resulted in loss of production of both peptides, demonstrating that they are processed in an eriB-dependent manner. Although precursors of ericins S and A show only 75% of identity, the matured lantibiotic-like peptides reveal highly similar physical properties; separation was only achieved after multistep, reversed-phase high-performance liquid chromatography. Based on Edman and peptidase degradation in combination with MALDI-TOF MS, for ericin S a subtilin-like, lanthionine-bridging pattern is supposed. For ericin A two C-terminal rings are different from the lanthionine pattern of subtilin. Due to only four amino acid exchanges, ericin S and subtilin revealed similar antibiotic activities as well as similar properties in response to heat and protease treatment. For ericin A only minor antibiotic activity was found.

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Creation of a Large Deletion Mutant of Campylobacter jejuni Reveals that the Lipooligosaccharide Gene Cluster Is Not Required for Viability

Journal of Bacteriology ◽

10.1128/jb.01397-08 ◽

2009 ◽

Vol 191 (7) ◽

pp. 2392-2399 ◽

Cited By ~ 7

Author(s):

Gemma L. Marsden ◽

Jianjun Li ◽

Paul H. Everest ◽

Andrew J. Lawson ◽

Julian M. Ketley

Keyword(s):

Gene Cluster ◽

Campylobacter Jejuni ◽

Deletion Mutant ◽

Natural Transformation ◽

Large Deletion ◽

Host Cells ◽

The Core ◽

Increased Sensitivity

ABSTRACT Deletion of the lipooligosaccharide biosynthesis region (Cj1132c to Cj1152c) from the genome of Campylobacter jejuni NCTC11168 shows that the core is not required for viability. The mutant was attenuated for growth and has increased sensitivity to antibiotics and detergents. Natural transformation and invasion of cultured host cells was abolished.

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