Barnes D E, Johnston L H, Kodama K, Tomkinson A E, Lasko D D, Lindahl T
Imperial Cancer Research Fund, Clare Hall Laboratories, South Mimms, Herts, United Kingdom.
Proc Natl Acad Sci U S A. 1990 Sep;87(17):6679-83. doi: 10.1073/pnas.87.17.6679.
Human cDNA clones encoding the major DNA ligase activity in proliferating cells, DNA ligase I, were isolated by two independent methods. In one approach, a human cDNA library was screened by hybridization with oligonucleotides deduced from partial amino acid sequence of purified bovine DNA ligase I. In an alternative approach, a human cDNA library was screened for functional expression of a polypeptide able to complement a cdc9 temperature-sensitive DNA ligase mutant of Saccharomyces cerevisiae. The sequence of an apparently full-length cDNA encodes a 102-kDa protein, indistinguishable in size from authentic human DNA ligase I. The deduced amino acid sequence of the human DNA ligase I cDNA is 40% homologous to the smaller DNA ligases of S. cerevisiae and Schizosaccharomyces pombe, homology being confined to the carboxyl-terminal regions of the respective proteins. Hybridization between the cloned sequences and mRNA and genomic DNA indicates that the human enzyme is transcribed from a single-copy gene on chromosome 19.
通过两种独立方法分离出了编码增殖细胞中主要DNA连接酶活性的人cDNA克隆，即DNA连接酶I。一种方法是，用从纯化的牛DNA连接酶I的部分氨基酸序列推导而来的寡核苷酸对人cDNA文库进行杂交筛选。另一种方法是，筛选人cDNA文库，以寻找能够互补酿酒酵母cdc9温度敏感型DNA连接酶突变体的多肽的功能表达。一个明显全长的cDNA序列编码一种102 kDa的蛋白质，其大小与天然人DNA连接酶I无法区分。人DNA连接酶I cDNA推导的氨基酸序列与酿酒酵母和裂殖酵母中较小的DNA连接酶有40%的同源性，同源性仅限于各自蛋白质的羧基末端区域。克隆序列与mRNA和基因组DNA之间的杂交表明，人酶是由19号染色体上的单拷贝基因转录而来的。
