Nagashima T, Yamamoto Y, Gomi K, Kitamoto K, Kumagai C
Shin Nihon Chemical Co., Ltd., Aichi, Japan.
Biosci Biotechnol Biochem. 1994 Jul;58(7):1292-6. doi: 10.1271/bbb.58.1292.
A high level production system for heterologous protein by cold culture of yeast transformants at 15 degrees C was developed. The yeast transformants, carrying a plasmid containing cDNA for Aspergillus oryzae alpha-amylase (Taka-amylase A) or human lysozyme synthetic DNA, were cultivated in a selective medium for 1 or 2 days until full growth at 30 degrees C. The yeast cells were harvested by centrifugation from the culture fluid and then were transferred to YPD medium. These inoculated broths were incubated for 2 days at 15 degrees C and then for another 2 days at 30 degrees C. By the cold culture method described above, higher amounts of Taka-amylase A (28.6 mg/liter) and human lysozyme (6.1 mg/liter) were produced by the yeast transformants compared to those by conventional methods. Heterologous protein productions using YEp, YCp, and YIp types of yeast expression vectors with ADH1 or GAPDH promoter by the cold culture method showed effective productivity of about 2-fold compared to those by the conventional method of culture at 30 degrees C. The high level production of heterologous protein by this method was not specific to the S. cerevisiae strains examined.
开发了一种通过在15℃下对酵母转化体进行冷培养来生产异源蛋白的高效生产系统。携带含有米曲霉α-淀粉酶（Taka-淀粉酶A）cDNA或人溶菌酶合成DNA的质粒的酵母转化体，在选择性培养基中于30℃培养1或2天直至完全生长。通过离心从培养液中收获酵母细胞，然后转移到YPD培养基中。将这些接种的肉汤在15℃下培养2天，然后在30℃下再培养2天。通过上述冷培养方法，与传统方法相比，酵母转化体产生了更高量的Taka-淀粉酶A（28.6毫克/升）和人溶菌酶（6.1毫克/升）。使用带有ADH1或GAPDH启动子的YEp、YCp和YIp类型酵母表达载体通过冷培养方法生产异源蛋白，与在30℃下的传统培养方法相比，显示出约2倍的有效生产率。通过这种方法高效生产异源蛋白并非所检测的酿酒酵母菌株所特有。
