외래 알파아밀라아제의 효모에서의 생산과 분비효율의 증진
Improvement of production and secretion rate of heterologous α-amylase in yeast
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In order to increase the production and secretion rate of heterologous mouse salivary α-amylase in yeast cell, various experiments were attempted. By replacing the native mouse secretion signal sequence with yeast invertase secretion signal sequence, the secretion rate was increased from 90% to 96%. A DNA fragment harbouring copper ionresistant gene(CUP_I) was inserted into the vector containing the α-amylase gene to ve pMS12(CUP_I). The transformant containing pMS12(CUP_I) showed 30% more α-amylase activity in the presence of 1 mMCuSO_4 in the culture medium, suggesting that the copy number of the vector containing the CUP_I in the yeast cell was increased by the exogenous copper ion and consequently the α-amylase in the same vector was amplified togeter with the vector. Several respiratory-deficient mutant strains were obtained by treating yeast cells with ethidium bromide, and the α-amylase activities of the respiratory-deficient mutant strains were 5∼8 times higher than those of parent strains. The composition of culture medium also affected the production rate of α-amylase. The 1% yeast extract and 2% peptone showed the highest activity of α-amylase. α-Amylase activity was also increased 3 times when the 0.015% (W/V) of 2-mercaptoethanol was added to the culture medium.