Expression, Glycosylation, and Secretion of an
            <i>Aspergillus</i>
            Glucoamylase by
            <i>Saccharomyces cerevisiae</i>

Innis, Michael A.; Holland, Michael J.; McCabe, Peter C.; Cole, G; Wittman, Vaughan; Tal, Rony; Watt, K W; Gelfand, David H.; Holland, J P; Meade, James H.

doi:10.1126/science.228.4695.21

Cited by 248 publications

(73 citation statements)

References 27 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Two popular methods can be used to detect cellular secretion, namely, secretome analysis and the glucoamylase assay (23). Although these methods are highly informative and convenient, 3 major problems arise when using them to detect unknown secretory pathways.…”

mentioning

confidence: 99%

Tracing Putative Trafficking of the Glycolytic Enzyme Enolase via SNARE-Driven Unconventional Secretion

et al. 2012

View full text Add to dashboard Cite

Glycolytic enzymes are cytosolic proteins, but they also play important extracellular roles in cell-cell communication and infection. We used Saccharomyces cerevisiae to analyze the secretory pathway of some of these enzymes, including enolase, phosphoglucose isomerase, triose phosphate isomerase, and fructose 1,6-bisphosphate aldolase. Enolase, phosphoglucose isomerase, and an N-terminal 28-amino-acid-long fragment of enolase were secreted in a sec23-independent manner. The enhanced green fluorescent protein (EGFP)-conjugated enolase fragment formed cellular foci, some of which were found at the cell periphery. Therefore, we speculated that an overview of the secretory pathway could be gained by investigating the colocalization of the enolase fragment with intracellular proteins. The DsRed-conjugated enolase fragment colocalized with membrane proteins at the cisGolgi complex, nucleus, endosome, and plasma membrane, but not the mitochondria. In addition, the secretion of full-length enolase was inhibited in a knockout mutant of the intracellular SNARE protein-coding gene TLG2. Our results suggest that enolase is secreted via a SNARE-dependent secretory pathway in S. cerevisiae.

show abstract

mentioning

confidence: 99%

Tracing Putative Trafficking of the Glycolytic Enzyme Enolase via SNARE-Driven Unconventional Secretion

et al. 2012

View full text Add to dashboard Cite

show abstract

“…It is more efficient for the fungi to use readily available carbon sources. Therefore, the production of the fungal glucoamylase is under carbon catabolite regulation (9,10). In a study on regulation of the glucoamylase from A. niger, Fowler et al (3) found that glucoamylase activity was observed when the fungus was grown on glucose.…”

Section: Discussionmentioning

confidence: 99%

Molecular Cloning, Characterization, and Differential Expression of a Glucoamylase Gene from the Basidiomycetous Fungus Lentinula edodes

Zhao

Chen

Kwan

2000

Appl Environ Microbiol

View full text Add to dashboard Cite

The complete nucleotide sequence of putative glucoamylase gene gla1 from the basidiomycetous fungus Lentinula edodes strain L54 is reported. The coding region of the genomic glucoamylase sequence, which is preceded by eukaryotic promoter elements CAAT and TATA, spans 2,076 bp. The gla1 gene sequence codes for a putative polypeptide of 571 amino acids and is interrupted by seven introns. The open reading frame sequence of the gla1 gene shows strong homology with those of other fungal glucoamylase genes and encodes a protein with an N-terminal catalytic domain and a C-terminal starch-binding domain. The similarity between the Gla1 protein and other fungal glucoamylases is from 45 to 61%, with the region of highest conservation found in catalytic domains and starch-binding domains. We compared the kinetics of glucoamylase activity and levels of gene expression in L. edodes strain L54 grown on different carbon sources (glucose, starch, cellulose, and potato extract) and in various developmental stages (mycelium growth, primordium appearance, and fruiting body formation). Quantitative reverse transcription PCR utilizing pairs of primers specific for gla1 gene expression shows that expression of gla1 was induced by starch and increased during the process of fruiting body formation, which indicates that glucoamylases may play an important role in the morphogenesis of the basidiomycetous fungus.Basidiomycetous fungus Lentinula edodes (Berk.) Pegler is the second most widely cultivated mushroom in the world. The cultivation of this fungus makes use of significant amounts of woody polysaccharides, and utilization of the complex polysaccharides is dependent on its ability to synthesize hydrolytic and oxidative enzymes which convert woody polysaccharides into low-molecular-weight compounds that can be absorbed and assimilated for nutrition. Starch is a polymer of glucose and is perhaps, next to cellulose, the most widely available polymeric glucoside made by plants (16,18). Starch is, therefore, available to fungi growing on plants or plant residues. Digestion of starch requires a complex of enzymes. Glucoamylases (1,4-␣-D-glucan glucohydrolases; EC 3.2.1.3) are enzymes that, among others, are believed to be important in the utilization of starch by the basidiomycetous fungus. Glucoamylases are exohydrolases, which catalyze the release of ␤-D-glucose units from the nonreducing ends of amylose, amylopectin, and other polysaccharides (18). Glucoamylase-encoding genes have been cloned from several fungi including Aspergillus awamori (6, 15), Aspergillus niger (1, 3), Aspergillus oryzae (8), Aspergillus terreus (5, 23), and Neurospora crassa (22).Although there have been many reports on glucoamylases in fungi, few studies on the production and regulation of glucoamylases in basidiomycetous fungi have been carried out. El-Zalaki and Hamza (2) studied five basidiomycetous fungus species for their ability to hydrolyze starch. L. edodes was found to be the most promising strain for amylase production. It was also reported that the ...

show abstract

“…For production of low-calorie beer in the brewing industry, it is of interest to use a recombinant strain of S. cerevisiae that secretes a glucoamylase whereby the larger oligomers (dextrins), which are formed from the partial hydrolysis of barley starch, are decomposed. Expression of a glucoamylase gene from Aspergillus awamori and secretion of the heterologous protein from S. cerevisiae were successfully demonstrated, but the transformed strain grew on dextrins at a lower rate than observed when glucoamylase was added externally to the medium (49). Coexpression of the STA2 gene of Saccharomyces diastaticus encoding a glucoamylase and an AMY1 gene encoding an ␣-amylase of Bacillus amyloliquefaciens was demonstrated to synergistically enhance starch degradation (131).…”

Section: Starch Utilizationmentioning

confidence: 96%

Metabolic Engineering of Saccharomyces cerevisiae

Østergaard

Olsson

Nielsen

2000

Microbiol Mol Biol Rev

399

213

View full text Add to dashboard Cite

SUMMARY Comprehensive knowledge regarding Saccharomyces cerevisiae has accumulated over time, and today S. cerevisiae serves as a widley used biotechnological production organism as well as a eukaryotic model system. The high transformation efficiency, in addition to the availability of the complete yeast genome sequence, has facilitated genetic manipulation of this microorganism, and new approaches are constantly being taken to metabolicially engineer this organism in order to suit specific needs. In this paper, strategies and concepts for metabolic engineering are discussed and several examples based upon selected studies involving S. cerevisiae are reviewed. The many different studies of metabolic engineering using this organism illustrate all the categories of this multidisciplinary field: extension of substrate range, improvements of producitivity and yield, elimination of byproduct formation, improvement of process performance, improvements of cellular properties, and extension of product range including heterologous protein production.

show abstract

Expression, Glycosylation, and Secretion of an Aspergillus Glucoamylase by Saccharomyces cerevisiae

Cited by 248 publications

References 27 publications

Tracing Putative Trafficking of the Glycolytic Enzyme Enolase via SNARE-Driven Unconventional Secretion

Tracing Putative Trafficking of the Glycolytic Enzyme Enolase via SNARE-Driven Unconventional Secretion

Molecular Cloning, Characterization, and Differential Expression of a Glucoamylase Gene from the Basidiomycetous Fungus Lentinula edodes

Metabolic Engineering of Saccharomyces cerevisiae

Contact Info

Product

Resources

About