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Crystal Structure of the Cancer Genomic DNA Mutator APOBEC3B Catalytic Domain
Author(s) -
Aihara Hideki,
Shi Ke,
Carpenter Michael A.,
Kurahashi Kayo,
Harris Reuben S.
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.1050.1
Subject(s) - active site , dna , mutagenesis , biology , binding site , nucleotide , chemistry , mutation , biochemistry , enzyme , gene
Background APOBEC3 family DNA cytosine deaminases provide overlapping defenses against pathogen infections as part of innate immunity. APOBEC3B is the only family member that is predominantly nuclear‐localized, and recent studies have demonstrated its major role in cancer mutagenesis. Methods We used the E. coli ‐based rifampicin resistance mutation assay and in vitro DNA deamination assay to guide the generation of a highly soluble and catalytically active APOBEC3B catalytic domain construct, and used X‐ray crystallography to analyze its structures. Results We have determined the first high‐resolution structures of APOBEC3B catalytic domain in multiple crystal forms. The structures revealed a tightly closed active site conformation, which suggest that the binding of a single‐stranded DNA substrate is regulated by adjacent flexible loops. Replacing one of these loops adjacent to the active site (loop 1) with the corresponding loop from the highly homologous cytoplasmic deaminase APOBEC3A generated a hyperactive enzyme, consistent with tight regulation of APOBEC3B activity. We also determined a nucleotide (dCMP)‐bound crystal structure that suggests a possible mode of APOBEC3B‐DNA interaction and informs a multi‐step model for the binding of single‐stranded DNA. Conclusion and Significance A comparison between our APOBEC3B structure and those of other cytoplasmic APOBEC3 members with more open active site conformations suggests that the active site accessibility of APOBEC3B may be regulated in a manner that maximizes innate antiviral activity and minimizes damages to genomic DNA. The novel nucleotide‐binding site on the surface of the APOBEC3B catalytic domain is accessible even when the active site is in the closed conformation, and thus it could serve as an initial contact point for a single‐stranded DNA substrate. Given its role in cancer mutagenesis and being a non‐essential enzyme, APOBEC3B is an attractive target for anti‐cancer drug development. Our high‐resolution structures provide a framework for further mechanistic studies and development of APOBEC3B inhibitors to suppress cancer evolution to minimize drug resistance and metastasis.