Glucoamylase gene, vldI, is linked to validamycin biosynthesis in Streptomyces hygroscopicus var. limoneus, and vldADEFG confers validamycin production in Streptomyces lividans, revealing the role of VldE in glucose attachment

Singh, Deepak; Kwon, Hyung‐Jin; Rajkarnikar, Arishma; Suh, Joo‐Won

doi:10.1016/j.gene.2007.02.015

Cited by 6 publications

(6 citation statements)

References 20 publications

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“…2000). Recently, it was shown that the knock‐out of a gene encoding a glucoamylase, vldI , reduced the yield of validamycin‐A, indicating that VldI contributes to validamycin‐A productivity by supplying glucose with the hydrolysis of 1,4‐ α ‐ d ‐glucan(s) (Singh et al. 2007).…”

Section: Discussionmentioning

confidence: 99%

Cloning of a gene encoding thermostable glucoamylase from Chaetomium thermophilum and its expression in Pichia pastoris

Chen

Zhang

Zhao

et al. 2007

Journal of Applied Microbiology

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Aims: Chaetomium thermophilum is a soil‐borne thermophilic fungus whose molecular biology is poorly understood. Only a few genes have been cloned from the Chaetomium genus. This study attempted to clone, to sequence and to express a thermostable glucoamylase gene of C. thermophilum. Methods and Results: First strand cDNA was prepared from total RNA isolated from C. thermophilum and the glucoamylase gene amplified by using PCR. Degenerate primers based on the N‐terminal sequences of the purified glucoamylase according to our previous works and a cDNA fragment encoding the glucoamylase gene was obtained through RT‐PCR. Using RACE‐PCR, full‐length cDNA of glucoamylase gene was cloned from C. thermophilum. The full‐length cDNA of the glucoamylase was 2016 bp and contained a 1797‐bp open reading frame encoding a protein glucoamylase precursor of 599 amino acid residues. The amino‐acid sequence from 31 to 45 corresponded to the N‐terminal sequence of the purified protein. The first 30 amino acids were presumed to be a signal peptide. The alignment results of the putative amino acid sequence showed the catalytic domain of the glucoamylase was high homology with the catalytic domains of the other glucoamylases. The C. thermophilum glucoamylase gene was expressed in Pichia pastoris, and the glucoamylase was secreted into the culture medium by the yeast in a functionally active form. The recombinant glucoamylase purified was a glycoprotein with a size of about 66 kDa, and exhibited optimum catalytic activity at pH 4·5–5·0 and 65°C. The enzyme was stable at 60°C, the enzyme activity kept 80% after 60 min incubation at 70°C. The half‐life was 40 and 10 min under incubation at 80 and 90°C respectively. Conclusions: A new thermostable glucoamylase gene of C. thermophilum was cloned, sequenced, overexpressed successfully in P. pastoris. Significance and Impact of the Study: Because of its thermostability and overexpression, this glucoamylase enzyme offers an interesting potential in saccharification steps in both starch enzymatic conversion and in alcohol production.

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Section: Discussionmentioning

confidence: 99%

Cloning of a gene encoding thermostable glucoamylase from Chaetomium thermophilum and its expression in Pichia pastoris

Chen

Zhang

Zhao

et al. 2007

Journal of Applied Microbiology

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“…The benzylated validamine and valienamine products were chromatographically separated from their keto-analogues, and the mixture was subjected to Cbz-protection to give 1-Cbz-2,3,4,7-tetrabenzylvalienamine ( 14) and 1-Cbz-2,3,4,7-tetrabenzylvalidamine (15). The compounds were then chromatographically separated, and complete deprotection of 15 by hydrogenolysis afforded validamine (16) in a quantitative yield.…”

Section: ' Resultsmentioning

confidence: 99%

“…Interestingly, VldE has previously been reported as a glycosyltransferase that catalyzes the glucosylation of validoxylamine A to give validamycin A. 16 However, a wealth of evidence is available to conclude that another glycosyltransferase, ValG (VldK), is responsible for this conversion. 2,17 Nevertheless, we explored the reported glycosyltransferase activity of VldE by incubating the enzyme with validoxylamine A and UDP-glucose and analyzing the product by ESIMS.…”

Section: ' Resultsmentioning

confidence: 99%

Pseudoglycosyltransferase Catalyzes Nonglycosidic C–N Coupling in Validamycin A Biosynthesis

et al. 2011

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Glycosyltransferases are ubiquitous in nature. They catalyze a glycosidic bond formation between sugar donors and sugar or non-sugar acceptors to produce oligo/polysaccharides, glycoproteins, glycolipids, glycosylated natural products, and other sugar-containing entities. However, a trehalose 6-phosphate synthase-like protein has been found to catalyze an unprecedented non-glycosidic C-N bond formation in the biosynthesis of the aminocyclitol antibiotic validamycin A. This dedicated ‘pseudoglycosyltransferase’ catalyzes a condensation between GDP-valienol and validamine 7-phosphate to give validoxylamine A 7′-phosphate with net retention of the ‘anomeric’ configuration of the donor cyclitol in the product. The enzyme operates in sequence with a phosphatase, which dephosphorylates validoxylamine A 7′-phosphate to validoxylamine A.

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“…Inactivation of vldI in Streptomyces hygroscopicus subsp. limoneus decreases validamycin production (Singh et al, 2007). VldI, as SlgC1 and SlgC2, lacks the CBM and is predicted to be intracellular.…”

Section: Figmentioning

confidence: 99%

Participation of putative glycoside hydrolases SlgC1 and SlgC2 in the biosynthesis of streptolydigin in Streptomyces lydicus

Gómez

Horna

Olano

et al. 2012

Microbial Biotechnology

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Summary Two genes of the streptolydigin gene cluster in Streptomyces lydicus cluster encode putative family 16 glycoside hydrolases. Both genes are expressed when streptolydigin is produced. Inactivation of these genes affects streptolydigin production when the microorganism is grown in minimal medium containing either glycerol or d‐glucans as carbon source. Streptolydigin yields in S. lydicus were increased by overexpression of either slgC1 or slgC2.

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Glucoamylase gene, vldI, is linked to validamycin biosynthesis in Streptomyces hygroscopicus var. limoneus, and vldADEFG confers validamycin production in Streptomyces lividans, revealing the role of VldE in glucose attachment

Cited by 6 publications

References 20 publications

Cloning of a gene encoding thermostable glucoamylase from Chaetomium thermophilum and its expression in Pichia pastoris

Cloning of a gene encoding thermostable glucoamylase from Chaetomium thermophilum and its expression in Pichia pastoris

Pseudoglycosyltransferase Catalyzes Nonglycosidic C–N Coupling in Validamycin A Biosynthesis

Participation of putative glycoside hydrolases SlgC1 and SlgC2 in the biosynthesis of streptolydigin in Streptomyces lydicus

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