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Title: 1.92 Angstrom Zinc-Free APOBEC3F Catalytic Domain Crystal Structure

Abstract

The APOBEC3 family of DNA cytosine deaminases is capable of restricting the replication of HIV-1 and other pathogens. Here, we report a 1.92 Å resolution crystal structure of the Vif-binding and catalytic domain of APOBEC3F (A3F). This structure is distinct from the previously published APOBEC and phylogenetically related deaminase structures, as it is the first without zinc in the active site. We determined an additional structure containing zinc in the same crystal form that allows direct comparison with the zinc-free structure. In the absence of zinc, the conserved active site residues that normally participate in zinc coordination show unique conformations, including a 90 degree rotation of His249 and disulfide bond formation between Cys280 and Cys283. We found that zinc coordination is influenced by pH, and treating the protein at low pH in crystallization buffer is sufficient to remove zinc. Zinc coordination and catalytic activity are reconstituted with the addition of zinc only in a reduced environment likely due to the two active site cysteines readily forming a disulfide bond when not coordinating zinc. We show that the enzyme is active in the presence of zinc and cobalt but not with other divalent metals. Furthermore, these results unexpectedly demonstrate that zincmore » is not required for the structural integrity of A3F and suggest that metal coordination may be a strategy for regulating the activity of A3F and related deaminases.« less

Authors:
 [1];  [1];  [1];  [1];  [1]
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  1. Univ. of Minnesota, Minneapolis, MN (United States)
Publication Date:
Research Org.:
Argonne National Laboratory (ANL), Argonne, IL (United States)
Sponsoring Org.:
USDOE; NIH NIGMS; NIH-ORIP HEI
OSTI Identifier:
1255304
Alternate Identifier(s):
OSTI ID: 1425518
Grant/Contract Number:  
AC02-06CH11357; GM095558; GM109770; P41 GM103403; S10 RR029205
Resource Type:
Accepted Manuscript
Journal Name:
Journal of Molecular Biology
Additional Journal Information:
Journal Volume: 428; Journal Issue: 11; Journal ID: ISSN 0022-2836
Publisher:
Elsevier
Country of Publication:
United States
Language:
ENGLISH
Subject:
59 BASIC BIOLOGICAL SCIENCES; cytosine deaminase; zinc-free and zinc-bound APOBEC3 structures; HIV-1 restriction factor; APOBEC3 regulation; X-ray crystallography

Citation Formats

Shaban, Nadine M., Shi, Ke, Li, Ming, Aihara, Hideki, and Harris, Reuben S. 1.92 Angstrom Zinc-Free APOBEC3F Catalytic Domain Crystal Structure. United States: N. p., 2016. Web. doi:10.1016/j.jmb.2016.04.026.
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Shaban, Nadine M., Shi, Ke, Li, Ming, Aihara, Hideki, & Harris, Reuben S. 1.92 Angstrom Zinc-Free APOBEC3F Catalytic Domain Crystal Structure. United States. https://doi.org/10.1016/j.jmb.2016.04.026
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Shaban, Nadine M., Shi, Ke, Li, Ming, Aihara, Hideki, and Harris, Reuben S. Sat . "1.92 Angstrom Zinc-Free APOBEC3F Catalytic Domain Crystal Structure". United States. https://doi.org/10.1016/j.jmb.2016.04.026. https://www.osti.gov/servlets/purl/1255304.
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@article{osti_1255304,
title = {1.92 Angstrom Zinc-Free APOBEC3F Catalytic Domain Crystal Structure},
author = {Shaban, Nadine M. and Shi, Ke and Li, Ming and Aihara, Hideki and Harris, Reuben S.},
abstractNote = {The APOBEC3 family of DNA cytosine deaminases is capable of restricting the replication of HIV-1 and other pathogens. Here, we report a 1.92 Å resolution crystal structure of the Vif-binding and catalytic domain of APOBEC3F (A3F). This structure is distinct from the previously published APOBEC and phylogenetically related deaminase structures, as it is the first without zinc in the active site. We determined an additional structure containing zinc in the same crystal form that allows direct comparison with the zinc-free structure. In the absence of zinc, the conserved active site residues that normally participate in zinc coordination show unique conformations, including a 90 degree rotation of His249 and disulfide bond formation between Cys280 and Cys283. We found that zinc coordination is influenced by pH, and treating the protein at low pH in crystallization buffer is sufficient to remove zinc. Zinc coordination and catalytic activity are reconstituted with the addition of zinc only in a reduced environment likely due to the two active site cysteines readily forming a disulfide bond when not coordinating zinc. We show that the enzyme is active in the presence of zinc and cobalt but not with other divalent metals. Furthermore, these results unexpectedly demonstrate that zinc is not required for the structural integrity of A3F and suggest that metal coordination may be a strategy for regulating the activity of A3F and related deaminases.},
doi = {10.1016/j.jmb.2016.04.026},
journal = {Journal of Molecular Biology},
number = 11,
volume = 428,
place = {United States},
year = {Sat Apr 30 00:00:00 EDT 2016},
month = {Sat Apr 30 00:00:00 EDT 2016}
}
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https://doi.org/10.1016/j.jmb.2016.04.026
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    Mechanism of Enhanced HIV Restriction by Virion Coencapsidated Cytidine Deaminases APOBEC3F and APOBEC3G
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    • Hou, Shurong; Silvas, Tania V.; Leidner, Florian
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    • DOI: 10.1021/acs.jctc.8b00545

    Computational Model and Dynamics of Monomeric Full-Length APOBEC3G
    journal, October 2017

    Structural basis for targeted DNA cytosine deamination and mutagenesis by APOBEC3A and APOBEC3B
    journal, December 2016

    • Shi, Ke; Carpenter, Michael A.; Banerjee, Surajit
    • Nature Structural & Molecular Biology, Vol. 24, Issue 2
    • DOI: 10.1038/nsmb.3344

    APOBEC3H structure reveals an unusual mechanism of interaction with duplex RNA
    journal, October 2017

    Understanding the structural basis of HIV-1 restriction by the full length double-domain APOBEC3G
    journal, January 2020

    Deamination hotspots among APOBEC3 family members are defined by both target site sequence context and ssDNA secondary structure
    journal, January 2020

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    • Nucleic Acids Research, Vol. 48, Issue 3
    • DOI: 10.1093/nar/gkz1164

    Apolipoprotein B mRNA editing enzyme catalytic polypeptide-like family genes activation and regulation during tumorigenesis
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    • Gao, Jianlong; Choudhry, Hani; Cao, Wei
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    APOBEC3B, a molecular driver of mutagenesis in human cancers
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