• Korean Journal of Food Science and Technology
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• v.16 no.3
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• pp.322-328
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• 1984

These experiments were conducted to purify the glucoamylase produced by Rhizopus oryzae. Two forms of glucoamylase (GI and GII) from Phizopus oryzae were purified by $(NH_2)_2SO_4$ fractionation, acetone fractionation and successive column chromatography on DEAE-cellulose and CM-cellulose. The specific activities of GI and GII toward soluble starch were 157.6 U/㎎. protein (37.5 fold of crude extract), and 164.7 U/㎎. protein (39.2 fold of curde extract), respectively, and the yields of them were 4.3% and 3.8%, respectively. The two purified enzymes have shown a single band by polyacrylamide disc gel electrophoresis and SDS-polyacrylamide gel electrophoresis. The protein bands of their electrophoresis gel were revealed to have glucoamylase activity by iodine staining and were proved to be glycoprotein by periodic acid Schiff's staining.

• Korean Journal of Food Science and Technology
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• v.32 no.4
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• pp.875-880
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• 2000

The production of glucoamylase and exopolygalacturonase from Cryptococcus laurentii Y-23 were investigated with the inducible substrates and mineral salts. Soluble starch induced only glucoamylase wherease pectin induced exopolygalacturonase as well as glucoamylase, and glucose did not induce glucoamylase whereas pectic acid induced a little amount of exopolygalacturonase. At the productions of two enzymes by inducible substrates for the 5 day-cultivation, the yeasts started log phase around 12 hours and mostly reached stationary phase around 36 hours. The best productivity of glucoamylase was observed with addition of soluble starch in the cultivation for 72 to 86 hours, and the high productivity of exopolygalacturonase was done by addition of both pectin and soluble starch in the cultivation for more than 72 hours. Without ammonium sulfate in the medium, however, cultural pH was so increased gradually that production of both enzymes were decreased and delayed as well. $Mn^{2+}$ increased both productivities of glucoamylase and exopolygalacturonase with 21% and 18%, respectively.

• Journal of Life Science
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• v.20 no.5
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• pp.683-690
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• 2010

The yeast strains of Saccharomyces diastaticus produce one of three isozymes of an extracellular glucoamylase I, II or III, a type of exo-enzyme which can hydrolyse starch to generate glucose molecules from non-reducing ends. These enzymes are encoded by the STA1, STA2 and STA3 genes. Another gene, sporulation-specific glucoamylase (SGA), also exists in the genus Saccharomyces which is very homologous to the STA genes. The SGA has been known to be produced in the cytosol during sporulation. However, we hypothesized that the SGA is capable of being secreted to the extracellular region because of about 20 hydrophobic amino acid residues at the N-terminus which can function as a signal peptide. We expressed the cloned SGA gene in S. diastaticus YIY345. In order to compare the biochemical properties of the extracellular glucoamylase and the SGA, the SGA was purified from the culture supernatant through ammonium sulfate precipitation, DEAE-Sephadex A-50, CM-Sephadex C-50 and Sephadex G-200 chromatography. The molecular weight of the intact SGA was estimated to be about 130 kDa by gel filtration chromatography with high performance liquid chromatography (HPLC) column. Sodium dedecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis showed it was composed of two heterogeneous subunits, 63 kDa and 68 kDa. The deglycosylation of the SGA generated a new 59 kDa band on the SDS-PAGE analysis, indicating that two subunits are glycosylated but the extent of glycosylation is different between them. The optimum pH and temperature of the SGA were 5.5 and $45^{\circ}C$, respectively, whereas those for the extracellular glucoamylase were 5.0 and $50^{\circ}C$. The SGA were more sensitive to heat and SDS than the extracellular glucoamylase.

• Microbiology and Biotechnology Letters
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• v.14 no.5
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• pp.365-369
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• 1986

The activity of glucoamylase and pullulanase, properties of glucoamylase and ethanol productivities of fusants were studied. Glucoamylase and pullulanase activity of fusants were higher than parents. The optimal pH and temperature of glucoamylase of fusants were very similar to the those produced by S. diastaticus. In alcohol fermentation. fermenting ability and fermentation rate of fusants were higher and faster than either of its parental strain. The maximum of alcohol yield in 15% of liquefied potato starch was 7.8% (v/v)

• Microbiology and Biotechnology Letters
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• v.16 no.3
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• pp.231-237
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• 1988

Several methods to produce ethanol from tapioca starch were examined. Among four methods tested, alcohol fermentation with extruded tapioca starch was the most effective, which alcohol yield was 460.5 f/ton. After 69hours reaction with Rhizopus sp. glucoamylase, 108.7mg/$m\ell$ of reducing sugar were produced from extruded tapioca and 43.8mg/$m\ell$ from raw tapioca starch. In alcohol fermentation with extruded tapioca, the high concentration of alcohol at early stage prevented bacterial contamination and the fermentation rate was increased due to the high saccharifying power of glucoamylase on the extruded starch, but extrusion temperature had no influence on the fermentability, Scanning electron microscopy showed that the extrusion process changed the structure of tapioca starch granule to more susceptible form to glucoamylase attack than the raw starch. And glucoamylase of Rhizopus sp. had stronger digestion activity on both extruded tapioca and raw tapioca starch than that of Aspergillus usamii.

• Journal of Microbiology and Biotechnology
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• v.11 no.1
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• pp.13-16
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• 2001

Maltose and soluble starch were used as secondary sources of carbon for glucoamylase production by Aspergillus awamori in solid state fermentation. During batch cultivation, maltose above 2.5%(w/w) repressed glucoamylase production, but, by adding either 2.5% (w/w) maltose or 1.25% (w/w) soluble starch to fed-batch cultivations, glucoamylase activity was increased by 15% and 170% over standard medium, respectively. The data showed that maltose is a weak inducer of glucoamylase production in solid stat fermentation.

• Microbiology and Biotechnology Letters
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• v.24 no.6
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• pp.712-716
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• 1996

The effects of non-ionic surfactants (Triton X-100 and Tween 80) on cloned glucoamylase production and secretion in recombinant Saccharomyces cerevisiae culture were studied. Even though the extracellular glucoamylase activity was increased by addition of Tween 80 due to the increase of the cell mass, Tween 80 did not play a role in the increase of glucoamylase secretion. On the addition of Triton X-100 addition, the secretion efficiency was increased while the cell growth was inhibited. Triton X-100 was added to the culture broth after 24 hr of culture to minimize the inhibition of the cell growth, and consequently the glucoamylase activity in the culture broth was increased by 12%.

• Journal of Microbiology and Biotechnology
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• v.25 no.2
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• pp.185-195
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• 2015

Glucoamylase is an important industrial enzyme. Glucoamylase production by industrial Aspergillus niger strain featured with two major problems: (i) empirical substrate feeding methods deteriorating the fermentation performance; and (ii) the high raw materials cost limiting the economics of the glucoamylase product with delegated specification. In this study, we first proposed a novel three-stage varied-rate substrate feeding strategy for efficient glucoamylase production in a 5 L bioreactor using the standard feeding medium, by comparing the changing patterns of the important physiological parameters such as DO, OUR, RQ, etc., when using different substrate feeding strategies. With this strategy, the glucoamylase activity and productivity reached higher levels of 11,000 U/ml and 84.6 U/ml/h, respectively. The performance enhancement in this case was beneficial from the following results: DO and OUR could be controlled at the higher levels (30%, 43.83 mmol/l/h), while RQ was maintained at a stable/lower level of 0.60 simultaneously throughout the fed-batch phase. Based on this three-stage varied-rate substrate feeding strategy, we further evaluated the economics of using alternative carbon sources, attempting to reduce the raw materials cost. The results revealed that cornstarch hydrolysate could be considered as the best carbon source to replace the standard and expensive feeding medium. In this case, the production cost of the glucoamylase with delegated specification (5,000 U/ml) could be saved by more than 61% while the product quality be ensured simultaneously. The proposed strategy showed application potential in improving the economics of industrial glucoamylase production.

• Journal of Mushroom
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• v.9 no.2
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• pp.53-58
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• 2011

In order to confirm the presence of putative glucoamylase gene in Tricholoma matsutake genome, the genomic DNA was prepared from T. matsutake NBRC30773 strain and was used as template to clone the glucoamylases gene (TmGlu1). We obtained the nucleotide sequence of TmGlu1 and its franking region. The coding region (from ATG to stop codon) is 2,186 bp. The locations of exons and introns were determined from the nucleotide sequences of 3'- and 5'-RACE PCR and RT-PCR products. On the other hand, to investigate the relationship between composition of medium and glucoamylase expression, we checked the expression level of glucoamylase gene by realtime reverse transcription PCR and measurement of glucoamylase enzyme activity. It was found that enzyme activity of glucoamylase was very low in different medium. Expression of glucoamylases gene appeared to not be affected by different carbon source.

• Korean Journal of Microbiology
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• v.28 no.3
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• pp.181-187
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• 1990

STA gene coding glucoamylase was introduced into haploid Saccharomyces cerevisiae SHY3 and polyploid Saccharomyces cerevisiae 54. We constructed the recombinant plasmid by substituting the promoter region of alcohol dehydrogenase isoenzyme I gene for that of STA gene to increase the expression of STA gene and found that the activity of glucoamylase was increased in transformants. The plasmid stability was improved remarkably when we got the STA gene into the plasmid which had centromere. The activity of glucoamylase and transformation frequency of it, however, was decreased because of low copy number. Industrial polyploid strain was transformed with the recombinant plasmid having the $2\mu$ origin of replication and STA gene. It produced more alcohol than host when fermented in liquefied starch media. The industrial strain, however, was not transformed with the autonomously replicating plasmid containing centromere.