title: On the solution conformation and dynamics of the HIV-1 viral infectivity factor.
abstract: Human immunodeficiency virus-1 (HIV-1) has evolved a cunning mechanism to circumvent the antiviral activity of the APOBEC3 family of host cell enzymes. HIV-1 Vif [viral (also called virion) infectivity factor], one of several HIV accessory proteins, targets APOBEC3 proteins for proteasomal degradation and downregulates their expression at the mRNA level. Despite the importance of Vif for HIV-1 infection, there is little conformational data on Vif alone or in complex with other cellular factors due to incompatibilities with many structural techniques and difficulties in producing suitable quantities of the protein for biophysical analysis. As an alternative, we have turned to hydrogen exchange mass spectrometry (HX MS), a conformational analysis method that is well suited for proteins that are difficult to study using X-ray crystallography and/or NMR. HX MS was used to probe the solution conformation of recombinant full-length HIV-1 Vif. Vif specifically interacted with the previously identified binding partner Hck and was able to cause kinase activation, suggesting that the Vif studied by HX MS retained a biochemically competent conformation relevant to Hck interaction. HX MS analysis of Vif alone revealed low deuteration levels in the N-terminal portion, indicating that this region contained structured or otherwise protected elements. In contrast, high deuteration levels in the C-terminal portion of Vif indicated that this region was likely unstructured in the absence of cellular interacting proteins. Several regions within Vif displayed conformational heterogeneity in solution, including the APOBEC3G/F binding site and the HCCH zinc finger. Taken together, these HX MS results provide new insights into the solution conformation of Vif.
authors.affiliation: Department of Chemistry and Chemical Biology and the Barnett Institute of Chemical and Biological Analysis, Northeastern University, Boston, MA 02115, USA.
authors.forename: Sean R
authors.initials: SR
authors.lastname: Marcsisin
authors.forename: Purushottam S
authors.initials: PS
authors.lastname: Narute
authors.forename: Lori A
authors.initials: LA
authors.lastname: Emert-Sedlak
authors.forename: Marek
authors.initials: M
authors.lastname: Kloczewiak
authors.forename: Thomas E
authors.initials: TE
authors.lastname: Smithgall
authors.forename: John R
authors.initials: JR
authors.lastname: Engen
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D055592:Biophysical Phenomena / Q000187:drug effects; D041961:Deuterium Exchange Measurement; D004789:Enzyme Activation / Q000187:drug effects; D015497:HIV-1 / Q000737:chemistry* / Q000187:drug effects; D006801:Humans; D013058:Mass Spectrometry; D008954:Models, Biological; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D051572:Proto-Oncogene Proteins c-hck / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012996:Solutions; D015032:Zinc / Q000494:pharmacology; D054320:vif Gene Products, Human Immunodeficiency Virus / Q000737:chemistry* / Q000378:metabolism
chemical_list: D010455:Peptides; D011994:Recombinant Proteins; D012996:Solutions; D054320:vif Gene Products, Human Immunodeficiency Virus; D051572:Proto-Oncogene Proteins c-hck; D015032:Zinc