Purification and structure of 3-methyladenine-DNA glycosylase I of Escherichia coli

K. Sakumi, Y. Nakabeppu, Y. Yamamoto, S. Kawabata, S. Iwanaga, M. Sekiguchi

研究成果: Contribution to journalArticle査読

32 被引用数 (Scopus)

抄録

We constructed a recombinant plasmid carrying a gene that suppresses tag mutation. To overproduce its gene product, a 0.8-kilobase DNA fragment which carries the gene was placed under the control of the lac promoter in pUC8. 3-Methyladenine-DNA glycosylase activity in cells carrying such plasmids (pCY5) was 450-fold higher than that of wild type strain, on exposure to isopropyl-β-D-thiogalactopyranoside. From an extract of such cells, the enzyme was purified to apparent physical homogeneity, and the amino acid composition and the amino-terminal amino acid sequence of the enzyme were determined. The data were in accord with nucleotide sequence of the gene, determined by the dideoxy method. It was deduced that 3-methyladenine-DNA glycosylase I comprises 187 amino acids and its molecular weight is 21,100, consistent with the value estimated from the sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the purified protein. Only 3-methyladenine was excised from methylated DNA by the purified glycosylase. These results show that the tag is the structural gene for 3-methyladenine-DNA glycosylase I.

本文言語英語
ページ(範囲)15761-15766
ページ数6
ジャーナルJournal of Biological Chemistry
261
33
出版ステータス出版済み - 1986

All Science Journal Classification (ASJC) codes

  • 生化学
  • 分子生物学
  • 細胞生物学

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