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"Purification and structure of 3-methyladenine-DNA glycosylase I of Escherichia coli." |
Sakumi K, Nakabeppu Y, Yamamoto Y, Kawabata S, Iwanaga S, Sekiguchi M |
Published Nov. 25, 1986 in J Biol Chem volume 261 .
Pubmed ID: 3536912
Abstract:
| 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-beta-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. |
This publication refers to following REPAIRtoire entries:
| Proteins |
Last modification of this entry: Oct. 6, 2010
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