CLONING AND NUCLEOTIDE-SEQUENCE OF THE KHS KILLER GENE OF SACCHAROMYCES-CEREVISIAE
CLONING AND NUCLEOTIDE-SEQUENCE OF THE KHS KILLER GENE OF SACCHAROMYCES-CEREVISIAE
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DOI：
10.1080/00021369.1991.10870924
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发表时间：
1991-08-01
期刊：
影响因子：
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通讯作者：
HARA, S
中科院分区：
文献类型：
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作者：
GOTO, K;FUKUDA, H;HARA, S
以酵母YEp13为载体，以大肠杆菌为宿主，从酿酒酵母115号基因组文库中克隆了KHS基因的一段5.3kbp的片段。用含有KHS基因的多拷贝载体将一株非致死酵母转化为致死菌株，转化子分泌的致死毒素是供体菌株115的3~4倍。通过凝胶过滤和柱层析，将KHS毒素从培养滤液中纯化了80倍。对KHS DNA中足以表达杀伤活性的一段2.8kbp的核苷酸序列进行了鉴定，发现了一个由2124bp组成的开放阅读框。对纯化的KHS毒素的开放阅读框和N端氨基酸序列进行了比较，推测KHS基因的多肽可能在分泌前的36Gln到37Ala之间被加工。
A 5.3-kbp fragment of the KHS gene was cloned from a genomic bank of Saccharomyces cerevisiae No. 115 constructed with an E. coli as the host and YEp13 as the vector. A non-killer yeast strain was transformed to a killer strain with the multi-copy vector containing the KHS gene, and the transformant could secrete 3-4 times more killer toxin into culture media than the donor, strain No. 115. The KHS toxin was purified 80-fold from the culture filtrate by gel filtration and column chromatography. The nucleotide sequence of a 2.8-kbp fragment of the KHS DNA that was enough for the expression of the killer activity was identified, and we found an open reading frame consisted of 2124bp. Comparison of the open reading frame and N-terminal amino acid sequence of purified KHS toxin, suggested that the presumed peptide from the KHS gene might be processed between 36Gln and 37Ala before secretion.