A Direct Interaction with RNA Dramatically Enhances the Catalytic Activity of the HIV-1 Protease In Vitro. Issue 14 (17th July 2015)

• Record Type: Journal Article
• Authors: Potempa, Marc; Nalivaika, Ellen; Ragland, Debra; Lee, Sook-Kyung; Schiffer, Celia A.; Swanstrom, Ronald
• Is Part Of: Journal of molecular biology. Volume 427:Issue 14(2015:Jul. 15)
• Page Start: 2360
• Page End: 2378
• Publication Date: 2015-07-17
• Subjects: HIV human immunodeficiency virus -- PFV prototype foamy virus -- MA matrix -- CA capsid -- NC nucleocapsid -- SP1 spacer peptide 1 -- SP2 spacer peptide 2 -- ssDNA single-stranded DNA -- NIH National Institutes of Health; Gag -- Maturation -- p15NC -- Protease -- Allosteric binding site; Molecular biology -- Periodicals; Biology -- Periodicals; Biochemistry -- Periodicals; Bacteriology -- Periodicals; Molecular Biology -- Periodicals; Biochemistry -- Periodicals; Biologie moléculaire -- Périodiques; Biologie -- Périodiques; Biochimie -- Périodiques; Moleculaire biologie; Biochemistry; Biology; Molecular biology; Periodicals; 572.805
• DOI: 10.1016/j.jmb.2015.05.007
• Languages: English
• ISSNs: 0022-2836

Abstract

Abstract: Though the steps of human immunodeficiency virus type 1 (HIV-1) virion maturation are well documented, the mechanisms regulating the proteolysis of the Gag and Gag-Pro-Pol polyproteins by the HIV-1 protease (PR) remain obscure. One proposed mechanism argues that the maturation intermediate p15NC must interact with RNA for efficient cleavage by the PR. We investigated this phenomenon and found that processing of multiple substrates by the HIV-1 PR was enhanced in the presence of RNA. The acceleration of proteolysis occurred independently from the substrate's ability to interact with nucleic acid, indicating that a direct interaction between substrate and RNA is not necessary for enhancement. Gel-shift assays demonstrated the HIV-1 PR is capable of interacting with nucleic acids, suggesting that RNA accelerates processing reactions by interacting with the PR rather than the substrate. All HIV-1 PRs examined have this ability; however, the HIV-2 PR does not interact with RNA and does not exhibit enhanced catalytic activity in the presence of RNA. No specific sequence or structure was required in the RNA for a productive interaction with the HIV-1 PR, which appears to be principally, though not exclusively, driven by electrostatic forces. For a peptide substrate, RNA increased the kinetic efficiency of the HIV-1 PR by an order of magnitude, affecting both turnover rate ( k cat ) and substrate affinity ( K m ). These results suggest that an allosteric binding site exists on the HIV-1 PR and that HIV-1 PR activity during maturation could be regulated in part by the juxtaposition of the enzyme with virion-packaged RNA. Graphical Abstract: Highlights: The mechanisms regulating HIV-1 protease (PR) activity are poorly understood. RNA accelerates cleavage of multiple substrates, regardless of RNA-binding ability. The HIV-1 PR can interact with nucleic acid to increase its catalytic efficiency. The interaction is primarily electrostatic, but size/structural determinants exist. Interactions between the HIV-1 PR and RNA could affect PR activity in the virion.