Determinants of sequence-specificity within human AID and APOBEC3G |
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| Authors: | Michael A Carpenter Erandi Rajagurubandara Priyanga Wijesinghe Ashok S Bhagwat |
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| Affiliation: | 1. Department of Health Sciences, University of Milano-Bicocca, via Cadore 48, 20900 Monza, MI, Italy;2. Department of Pharmacological and Biomolecular Sciences, University of Milan, via Trentacoste 2, 20134 Milan, Italy;1. National Institute for Infectious Diseases “L. Spallanzani”, Rome, Italy;2. University of Rome “Tor Vergata”, Department of Biology, Rome, Italy;1. State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zu Chong Zhi Road, Shanghai 201203, China;2. Department of Physiology, Norman Bethune Medical College, Jilin University, Changchun 130021, China;3. School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing 210046, China;4. Phamacology department, China Pharmaceutical University, Nanjing 211198, China;5. Alcohol & Drug Abuse Research Center, McLean Hospital, Harvard Medical School, MA 02478, USA;3. Department of Biology, University of Western Ontario, London, Ontario N6A 5B7, Canada;4. Department of Biological Chemistry, School of Medicine, University of California Irvine, Irvine, California 92697 |
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| Abstract: | Human APOBEC3G (A3G) and activation-induced deaminase (AID) belong to a family of DNA-cytosine deaminases. While A3G targets the last C in a run of C's, AID targets C in the consensus sequence WRC (W is A or T and R is a purine). Guided by the structures of the A3G carboxyl-terminal catalytic domain (A3G-CTD), we identified two potential regions (region 1 and region 2) that may interact with DNA and swapped the corresponding regions between a variant of A3G-CTD and AID. The resulting hybrids were expressed in Escherichia coli and two different genetic assays and a biochemical assay were used to determine the sequence selectivity of the hybrids in promoting C to T mutations. The results show that while the 10 amino acid region 2 of A3G was its principal sequence-specificity determinant, region 1 of A3G enhanced the target cytosine preference conferred by region 2. In contrast, neither of the two regions in AID individually or in combination were sufficient to confer the DNA sequence preference of this protein upon A3G. Instead, introduction of AID sequences in A3G relaxed the sequence-specificity of the latter protein. Our results show that the sequence selectivity of APOBEC family of enzymes is determined by at least two separate sequence segments and there may be additional regions of the protein involved in DNA sequence recognition. |
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