© 1992 Oxford University Press
research-article |
Specific amino acid sequences required for O6-methylguanine-DNA methyltransferase activity: analyses of three residues at or near the methyl acceptor site
Department of Biochemistry, Faculty of Medicine and the Medical Institute of Bioregulation, Kyushu University Fukuoka 812, Japan
1To whom correspondence should be addressed
O6-Methylguanine-DNA methyltransferase plays an important role in preventing tumor induction. To elucidate the significance of a highly conserved amino acid sequence of methyltransferase protein, amino acid substitutions were introduced by site-directed mutagenesis of cloned cDNA for human methyltransferase and the activity and stability of mutant forms of enzyme were examined. When cysteine-145, to which the methyl transfer occurs, was replaced by other amino acids, all of the mutants isolated showed the methyl-transferase-negative phenotype. From one of the negative mutants, methyltransferase-positive revertants were isolated, all of which carried codons for cysteine. Thus the cysteine residue is essential for acceptance of the methyl group and cannot be replaced by other amino acids. Using this negative and positive selectionprocedure, analyses were extended to other residues near the acceptor site. At the histidine-146 site, four substitutions (phenyialanine, methionine, asparagine and glutamine) exhibited the positive phenotype but the levels of methyltransferaseactivity in these mutants were low. With valine-148 substitutions there were six types of positive revertants, among which mutants carrying isoleucine, cysteine and alanine showed significantly high levels of methyltransferase activity. Some mutant forms of cDNA were expressed in methyltransferase-defkient human cells, and the results obtained with Escherichia coli cells were confirmed.
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  Y. Zhong, Y. Huang, Y. Huang, T. Zhang, C. Ma, S. Zhang, W. Fan, H. Chen, J. Qian, and D. Lu Effects of O6-methylguanine-DNA methyltransferase (MGMT) polymorphisms on cancer: a meta-analysis Mutagenesis, November 5, 2009; (2009) gep050v1. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Hazra, S. Chanock, E. Giovannucci, D. G. Cox, T. Niu, C. Fuchs, W. C. Willett, and D. J. Hunter Large-Scale Evaluation of Genetic Variants in Candidate Genes for Colorectal Cancer Risk in the Nurses' Health Study and the Health Professionals' Follow-up Study Cancer Epidemiol. Biomarkers Prev., February 1, 2008; 17(2): 311 - 319. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Huang, F. Ye, H. Chen, W. Lu, and X. Xie Amino acid substitution polymorphisms of the DNA repair gene MGMT and the susceptibility to cervical carcinoma Carcinogenesis, June 1, 2007; 28(6): 1314 - 1322. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Shen, M. B. Terry, M. D. Gammon, M. M. Gaudet, S. L. Teitelbaum, S. M. Eng, S. K. Sagiv, A. I. Neugut, and R. M. Santella MGMT genotype modulates the associations between cigarette smoking, dietary antioxidants and breast cancer risk Carcinogenesis, December 1, 2005; 26(12): 2131 - 2137. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S Halford, A Rowan, E Sawyer, I Talbot, and I Tomlinson O6-methylguanine methyltransferase in colorectal cancers: detection of mutations, loss of expression, and weak association with G:C>A:T transitions Gut, June 1, 2005; 54(6): 797 - 802. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Iijima, J. Grant, K. McElroy, V. Fyfe, T. Preston, and K. E. L. McColl Novel mechanism of nitrosative stress from dietary nitrate with relevance to gastro-oesophageal junction cancers Carcinogenesis, December 1, 2003; 24(12): 1951 - 1960. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. Liu, S. Schwartz, B. M. Davis, and S. L. Gerson Chemotherapy-induced O6-Benzylguanine-resistant Alkyltransferase Mutations in Mismatch-deficient Colon Cancer Cancer Res., June 1, 2002; 62(11): 3070 - 3076. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Jaiswal, N. F. LaRusso, L. J. Burgart, and G. J. Gores Inflammatory Cytokines Induce DNA damage and Inhibit DNA repair in Cholangiocarcinoma Cells by a Nitric Oxide-dependent Mechanism Cancer Res., January 1, 2000; 60(1): 184 - 190. [Abstract] [Full Text]
|
|