Formation of transient dimers by a retroviral protease.
Retroviral proteases have been shown previously to be only active as homodimers. They are essential to form the separate and active proteins from the viral precursors. Spumaretroviruses produce separate precursors for Gag and Pol, rather than a Gag and a Gag-Pol precursor. Nevertheless, processing of Pol into a PR (protease)-RT (reverse transcriptase) and integrase is essential in order to obtain infectious viral particles. We showed recently that the PR-RT from a simian foamy virus, as well as the separate PRshort (protease) domain, exhibit proteolytic activities, although only monomeric forms could be detected. In the present study, we demonstrate that PRshort and PR-RT can be inhibited by the putative dimerization inhibitor cholic acid. Various other inhibitors, including darunavir and tipranavir, known to prevent HIV-1 PR dimerization in cells, had no effect on foamy virus protease in vitro. 1H-15N HSQC (heteronuclear single quantum coherence) NMR analysis of PRshort indicates that cholic acid binds in the proposed PRshort dimerization interface and appears to impair formation of the correct dimer. NMR analysis by paramagnetic relaxation enhancement resulted in elevated transverse relaxation rates of those amino acids predicted to participate in dimer formation. Our results suggest transient PRshort homodimers are formed under native conditions but are only present as a minor transient species, which is not detectable by traditional methods.
Hartl
Maximilian J
MJ
Universität Bayreuth, Lehrstuhl für Struktur und Chemie der Biopolymere and Research Centre for Biomacromolecules, Germany.
Schweimer
Kristian
K
Reger
Martin H
MH
Schwarzinger
Stephan
S
Bodem
Jochen
J
Rösch
Paul
P
Wöhrl
Birgitta M
BM
D001665:Binding Sites; D002460:Cell Line; D019826:Cholic Acid / Q000494:pharmacology; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D010447:Peptide Hydrolases / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D055503:Protein Multimerization / Q000187:drug effects; D012190:Retroviridae / Q000201:enzymology; D053150:Simian foamy virus / Q000201:enzymology
D010447:Peptide Hydrolases; D019826:Cholic Acid