Cloning and expression of the Saccharomycopsis fibuligera glucoamylase gene in Saccharomyces cerevisiae

Yamashita, Ichiro; Itoh, Tetsuya; Fukui, Sakuzô

doi:10.1007/bf00249943

Cited by 8 publications

(15 citation statements)

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“…Cells of S. fibuligera HUT7212 (30) were cultured in 100 ml of YPS medium (1% yeast extract, 2% polypeptone, and 3% soluble starch) to an optical density at 660 nm of 1.0, harvested, and washed with sterile water. RNA was extracted from the cells as described by Jensen et al (9).…”

Section: Methodsmentioning

confidence: 99%

“…Recently, we cloned the glucoamylase gene GLUI from a genomic library of S. fibuligera by screening Saccharomyces cerevisiae transformants capable of starch fermentation (30). The transformed cells secrete large amounts of glucoamylase into the growth medium such that the protein is shown to be only one major band after acrylamide gel electrophoresis of the culture fluid.…”

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confidence: 99%

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Nucleotide sequence of the glucoamylase gene GLU1 in the yeast Saccharomycopsis fibuligera

Itoh¹,

Ohtsuki²,

Yamashita³

et al. 1987

J Bacteriol

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The complete nucleotide sequence of the glucoamylase gene GLUI from the yeast Saccharomycopsisfibuligera has been determined. The GLUI DNA hybridized to a polyadenylated RNA of 2.1 kilobases. A single open reading frame codes for a 519-amino-acid protein which contains four potential N-glycosylation sites. The putative precursor begins with a hydrophobic segment that presumably acts as a signal sequence for secretion. Glucoamylase was purified from a culture fluid of the yeast Saccharomyces cerevisiae which had been transformed with a plasmid carrying GLUI. The molecular weight of the protein was 57,000 by both gel filtration and acrylamide gel electrophoresis. The protein was glycosylated with asparagine-linked glycosides whose molecular weight was 2,000. The amino-terminal sequence of the protein began from the 28th amino acid residue from the first methionine of the putative precursor. The amino acid composition of the purified protein matched the predicted amino acid composition. These results confirmed that GLUI encodes glucoamylase. A comparison of the amino acid sequence of glucoamylases from several fungi and yeast shows five highly conserved regions. One homology region is absent from the yeast enzyme and so may not be essential to glucoamylase function.

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

Nucleotide sequence of the glucoamylase gene GLU1 in the yeast Saccharomycopsis fibuligera

Itoh¹,

Ohtsuki²,

Yamashita³

et al. 1987

J Bacteriol

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“…cerevisiae has been a popular choice for heterelogous expression of proteins (48) and tested to express α-amylase and glucoamylase from S. fibuligera strain HUT 7212 (49,50). S.…”

Section: Amy Recombinant Behaviour On Purification In Chromatography mentioning

confidence: 99%

Chromatography as the Major Tool in the Identification and the Structure-Function Relationship Study of Amylolytic Enzymes from Saccharomycopsis Fibuligera R64

Ismaya¹,

Hasan²,

Subroto³

et al. 2012

Chromatography - The Most Versatile Method of Chemical Analysis

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“…GLU1 and GLA1 encode glucoamylase from Sacfi strain HUT7212 (Yamashita et al 1985) and KZ (Hostinova et al 1991), respectively. Intriguingly, while the sequence alignment of their gene products shows a high homology (Hostinova 2002), the GLU1 and GLA1 exhibit different characteristics.…”

Section: Introductionmentioning

confidence: 99%

Biochemical characterization of a glucoamylase from Saccharomycopsis fibuligera R64

et al. 2010

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Glucoamylase from the yeast Saccharomycopsis fibuligera R64 (GLL1) has successfully been purified and characterized. The molecular mass of the enzyme was 56,583 Da as determined by mass spectrometry. The purified enzyme demonstrated optimum activity in the pH range of 5.6-6.4 and at 50• C. The activity of the enzyme was inhibited by acarbose with the IC50 value of 5 µM. GLL1 shares high amino acid sequence identity with GLU1 and GLA1, which are Saccharomycopsis fibuligera glucoamylases from the strains HUT7212 and KZ, respectively. The properties of GLL1, however, resemble that of GLU1. The elucidation of the primary structure of GLL1 contributes to the explanation of this finding.

show abstract

Cloning and expression of the Saccharomycopsis fibuligera glucoamylase gene in Saccharomyces cerevisiae

Cited by 8 publications

References 11 publications

Nucleotide sequence of the glucoamylase gene GLU1 in the yeast Saccharomycopsis fibuligera

Nucleotide sequence of the glucoamylase gene GLU1 in the yeast Saccharomycopsis fibuligera

Chromatography as the Major Tool in the Identification and the Structure-Function Relationship Study of Amylolytic Enzymes from Saccharomycopsis Fibuligera R64

Biochemical characterization of a glucoamylase from Saccharomycopsis fibuligera R64

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