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KMID : 0364819990350040315
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Korean Journal of Microbiology
1999 Volume.35 No. 4 p.315 ~ p.321
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Characterization of α-amylase Producing Hybrid Constructed between Saccharomycopsis and Saccharomyces
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양영기:Yang Young-Ki
문명님:Moon Myeng-Nim/임채영:Lim Chae-Young/이영하:Rhee Young-Ha/김정호:Kim Jeong-Ho/:Glenn Chambliss
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Abstract
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This study was been performed to develope a years strain having high α-amylase
production ability using nuclear transfer method. Hybrids formed between the strains of
Saccharomyces fiburigera KCTC 7393 and Saccharomyces cerevisiae KCTC 7049 (tyr-,
ura-) were obtained by nuclear transfer technique. Nuclei isolated from the wild type S.
fiburigera strain were transfered into auxotrophic mutants S. cerevisiae and selected the
hybrids showing an increased starch degrading capability were selected (MN-16). This
transformant grew best and produced maximal α-amylase activity on the medium
containing 2% (V/V) soluble starch. α-Amylase from MN-16 was purified
electrophoretically homogenety and its properties were investigated. The enzyme was
purfied about 1.6 fold with an overall yield 9.7% from the culture medium by ammonium
sulfate fractionation. DEAE-Sephacel column chromatography, and Sephacryl S-200
column chromatography. The purified enzyme showed a single band on
SDS-polyacrylamide gel electrophoresis. The molecular weight of the α-amylase was
estimated to be 53,000 daltons by SDS-PAGE and by gel permeation chromatography on
Sephacryl S-200. The purified enzyme showed the maximum activity at pH 5.5 and 4
0℃. The km value for soluble starch was 2.5 mg/ml. The enzyme activity increased in
the presence of Ca2+, Co2+, EDTA, Mg2+, and
Zn2+, but inhibited by Cu2+, Fe2+, and
Ni2+.
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KEYWORD
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α-amylase, chromatography, characterization, Saccharomycopsis, Sacchromyces,
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