期刊
JOURNAL OF BIOLOGICAL CHEMISTRY
卷 287, 期 19, 页码 15826-15835出版社
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M112.342790
关键词
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资金
- National Institutes of Health [GM085116, ES13192, GM2142]
- National Science Foundation [MCB-0747285]
- Direct For Biological Sciences
- Div Of Molecular and Cellular Bioscience [0747285] Funding Source: National Science Foundation
APOBEC3G (Apo3G) is a single-stranded (ss) DNA cytosine deaminase that eliminates HIV-1 infectivity by converting C -> U in numerous small target motifs on the minus viral cDNA. Apo3G deaminates linear ssDNA in vitro with pronounced spatial asymmetry favoring the 3' -> 5' direction. A similar polarity observed in vivo is believed responsible for initiating localized C -> T mutational gradients that inactivate the virus. When compared with double-stranded (ds) DNA scanning enzymes, e.g. DNA glycosylases that excise rare aberrant bases, there is a paucity of mechanistic studies on ssDNA scanning enzymes. Here, we investigate ssDNA scanning and motif-targeting mechanisms for Apo3G using single molecule Forster resonance energy transfer. We address the specific issue of deamination asymmetry within the general context of ssDNA scanning mechanisms and show that Apo3G scanning trajectories, ssDNA contraction, and deamination efficiencies depend on motif sequence, location, and ionic strength. Notably, we observe the presence of bidirectional quasi-localized scanning of Apo3G occurring proximal to a 5' hot motif, a motif-dependent DNA contraction greatest for 5' hot > 3' hot > 5' cold motifs, and diminished mobility at low salt. We discuss the single molecule Forster resonance energy transfer data in terms of a model in which deamination polarity occurs as a consequence of Apo3G binding to ssDNA in two orientations, one that is catalytically favorable, with the other disfavorable.
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