Crystal Structure of a Soluble APOBEC3G Variant Suggests ssDNA to Bind in a Channel that Extends between the Two Domains
Delviks-Frankenberry, Krista A.
Sierra Rodriguez, Christina
Yilmaz, Nese Kurt
Pathak, Vinay K.
Schiffer, Celia A.
UMass Chan AffiliationsSchiffer Lab
Department of Biochemistry and Molecular Pharmacology
Document TypeJournal Article
Enzymes and Coenzymes
Medicinal Chemistry and Pharmaceutics
MetadataShow full item record
AbstractAPOBEC3G (A3G) is a single-stranded DNA (ssDNA) cytosine deaminase that can restrict HIV-1 infection by mutating the viral genome. A3G consists of a non-catalytic N-terminal domain (NTD) and a catalytic C-terminal domain (CTD) connected by a short linker. While the CTD catalyzes cytosine deamination, the NTD is believed to provide additional affinity for ssDNA. Structures of both A3G domains have been solved individually; however, a full-length A3G structure has been challenging. Recently, crystal structures of full-length rhesus macaque A3G variants were solved which suggested dimerization mechanisms and RNA binding surfaces, whereas the dimerization appeared to compromise catalytic activity. We determined the crystal structure of a soluble variant of human A3G (sA3G) at 2.5 A and from these data generated a model structure of wild-type A3G. This model demonstrated that the NTD was rotated 90 degrees relative to the CTD along the major axis of the molecule, an orientation that forms a positively charged channel connected to the CTD catalytic site, consisting of NTD loop-1 and CTD loop-3. Structure-based mutations, in vitro deamination and DNA binding assays, and HIV-1 restriction assays identify R24, located in the NTD loop-1, as essential to a critical interaction with ssDNA. Furthermore, sA3G was shown to bind a deoxy-cytidine dinucleotide near the catalytic Zn(2+), yet not in the catalytic position, where the interactions between deoxy-cytidines and CTD loop-1 and loop-7 residues were different from those formed with substrate. These new interactions suggest a mechanism explaining why A3G exhibits a 3' to 5' directional preference in processive deamination.
Maiti A, Myint W, Delviks-Frankenberry KA, Hou S, Kanai T, Balachandran V, Sierra Rodriguez C, Tripathi R, Kurt Yilmaz N, Pathak VK, Schiffer CA, Matsuo H. Crystal Structure of a Soluble APOBEC3G Variant Suggests ssDNA to Bind in a Channel that Extends between the Two Domains. J Mol Biol. 2020 Nov 20;432(23):6042-6060. doi: 10.1016/j.jmb.2020.10.020. Epub 2020 Oct 22. PMID: 33098858; PMCID: PMC7771068. Link to article on publisher's site