title: Amide hydrogen exchange in HIV-1 subtype B and C proteases--insights into reduced drug susceptibility and dimer stability.
abstract: Since its identification, HIV has continued to have a detrimental impact on the lives of millions of people throughout the world. The protease of HIV is a major target in antiviral treatment. The South African HIV-1 subtype C (C-SA) protease displays weaker binding affinity for some clinically approved protease inhibitors in comparison with the HIV-1 subtype B protease. The heavy HIV burden in sub-Saharan Africa, where subtype C HIV-1 predominates, makes this disparity a topic of great interest. In light of this, the enzyme activity and affinity of protease inhibitors for the subtype B and C-SA proteases were determined. The relative vitality, indicating the selective advantage of polymorphisms, of the C-SA protease relative to the subtype B protease in the presence of ritonavir and darunavir was four- and tenfold greater, respectively. Dynamic differences that contribute to the reduced drug susceptibility of the C-SA protease were investigated by performing hydrogen-deuterium exchange/mass spectrometry (HDX/MS) on unbound subtype B and C-SA proteases. The reduced propensity to form the E35-R57 salt bridge, and alterations in the hydrophobic core of the C-SA protease, are proposed to affect the anchoring of the flexible flaps, resulting in an increased proportion of the fully open flap conformation. HDX/MS data suggested that the N-terminus of both proteases is less stable than the C-terminus of the proteases, thus explaining the increased efficacy of dimerization inhibitors targeted toward the C-terminus of HIV proteases. As far as we are aware, this is the first report on assessment of HIV protease dynamics using HDX/MS.
authors.affiliation: Protein Structure-Function Research Unit, School of Molecular and Cell Biology, University of the Witwatersrand, Johannesburg, South Africa.
authors.forename: Previn
authors.initials: P
authors.lastname: Naicker
authors.forename: Stoyan
authors.initials: S
authors.lastname: Stoychev
authors.forename: Heini W
authors.initials: HW
authors.lastname: Dirr
authors.forename: Yasien
authors.initials: Y
authors.lastname: Sayed
mesh_terms: D000577:Amides / Q000378:metabolism*; D000595:Amino Acid Sequence; D019281:Dimerization; D016333:HIV Protease / Q000737:chemistry / Q000378:metabolism*; D015497:HIV-1 / Q000201:enzymology*; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid
chemical_list: D000577:Amides; D016333:HIV Protease; C514803:p16 protease, Human immunodeficiency virus 1
keywords: dimer interface; drug binding; flap conformations; hydrogen-deuterium exchange; protein dynamics