Cloning,sequencing and expression of the uvrA gene from an extremely thermophilic bacterium,Thermus thermophilus HB8 |
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| Affiliation: | 1. International Co-operation Laboratory on Signal Transduction, Eastern Hepatobiliary Surgery Institute, Second Military Medical University, Shanghai 200438, China;2. National Center for Liver Cancer, Shanghai, China;3. Qidong Liver Cancer Institute and Nantong University Liver Cancer Institute, QiDong 226200, Jiangsu, China;4. MOE Key Laboratory of Environment and Health, School of Public Health, Tongji Medical College, Huazhong University of Science and Technology, 13 Hangkong Rd, Wuhan 430030, China;5. State Key Lab of Genetic Engineering and MOE Key Lab of Contemporary Anthropology, TaiZhou Institute of Health Sciences and School of Life Sciences, Collaborative Innovation Center of Genetics and Development, Fudan University, Shanghai, China;6. State Key Lab Breeding Base of Xinjiang Major Diseases Research, First Affiliated Hospital of Xinjiang Medical University, Xinjiang 830054, China;7. Department of Epidemiology, Second Military Medical University, Shanghai 200433, China;8. Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai 200438, China;9. Jiangsu Key Laboratory of Molecular and Translational Cancer Research, Cancer Hospital of Jiangsu Province, Nanjing, China;10. Laboratory of Metabolism, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA;11. National Laboratory for Oncogenes and Related Genes, Cancer Institute, RenJi Hospital, Shanghai Jiao Tong University, Shanghai 200441, China;1. Department of Chemistry and Biochemistry, City College of New York, New York, NY 10031, USA;2. PhD Program in Biochemistry, The Graduate Center of the City University of New York, New York, NY 10016, USA;3. Department of Biophysics and Biophysical Chemistry, Johns Hopkins School of Medicine, Baltimore, MD 21205, USA;4. PhD Program in Biology, The Graduate Center of the City University of New York, New York, NY 10016, USA;5. PhD Program in Chemistry, The Graduate Center of the City University of New York, New York, NY 10016, USA |
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| Abstract: | One of the most important DNA repair systems is the nucleotide (nt) excision repair system. The uvrA gene, which plays an essential role in the prokaryotic excision repair system, was cloned from an extremely thermophilic eubacterium, Thermus thermophilus (Tt) HB8, and its nt sequence was determined. In the amino acid (aa) sequence of Tt UvrA, a characteristic duplicated structure, two nt-binding consensus sequences (Walker's A-type motif) and two zinc finger DNA-binding motifs were found. The aa sequence showed 73% homology with that of Escherichia coli (Ec). These features suggest that Tt has the same excision repair system as Ec. Upon comparison of the Tt and Ec UvrA, some characteristic aa substitutions were found. The numbers of Arg and Pro residues were increased (from 66 to 81 and from 47 to 55, respectively), and the numbers of Asn and Met residues were decreased (from 33 to 18 and from 18 to 11, respectively) in Tt. The Tt uvrA gene was expressed in Ec under control of the lac promoter. Purified UvrA was stable up to 80°C (at neutral pH) and at pH 2–11 (at 25°C) |
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