HIV restriction factor APOBEC3G binds in multiple steps and conformations to search and deaminate single-stranded DNA
- Northeastern University, United States
- University of Saskatchewan, Canada
- Ohio State University, United States
Abstract
APOBEC3G (A3G), an enzyme expressed in primates with the potential to inhibit human immunodeficiency virus type 1 (HIV-1) infectivity, is a single-stranded DNA (ssDNA) deoxycytidine deaminase with two domains, a catalytically active, weakly ssDNA binding C-terminal domain (CTD) and a catalytically inactive, strongly ssDNA binding N-terminal domain (NTD). Using optical tweezers, we measure A3G binding a single, long ssDNA substrate under various applied forces to characterize the binding interaction. A3G binds ssDNA in multiple steps and in two distinct conformations, distinguished by degree of ssDNA contraction. A3G stabilizes formation of ssDNA loops, an ability inhibited by A3G oligomerization. Our data suggests A3G securely binds ssDNA through the NTD, while the CTD samples and potentially deaminates the substrate. Oligomerization of A3G stabilizes ssDNA binding but inhibits the CTD's search function. These processes explain A3G's ability to efficiently deaminate numerous sites across a 10,000 base viral genome during the reverse transcription process.
Data availability
Source data files have been supplied for figures 2, 2 supplement, 3, 4, and 5. Additionally, custom written Matlab and Lab windows code is supplied with this manuscript.
Article and author information
Author details
Funding
National Institute of General Medical Sciences (GM072462)
- Michael Morse
- Ioulia Rouzina
- Mark C Williams
Canadian Institutes of Health Research (MOP137090)
- Yuqing Feng
- Linda Chelico
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2019, Morse et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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HIV restriction factor APOBEC3G binds in multiple steps and conformations to search and deaminate single-stranded DNA
eLife 8:e52649.
https://doi.org/10.7554/eLife.52649
