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Acetylation regulates cyclophilin A catalysis, immunosuppression and HIV isomerization.

ABSTRACT: Cyclophilin A (CypA) is a ubiquitous cis-trans prolyl isomerase with key roles in immunity and viral infection. CypA suppresses T-cell activation through cyclosporine complexation and is required for effective HIV-1 replication in host cells. We show that CypA is acetylated in diverse human cell lines and use a synthetically evolved acetyllysyl-tRNA synthetase/tRNA(CUA) pair to produce recombinant acetylated CypA in Escherichia coli. We determined atomic-resolution structures of acetylated CypA and its complexes with cyclosporine and HIV-1 capsid. Acetylation markedly inhibited CypA catalysis of cis to trans isomerization and stabilized cis rather than trans forms of the HIV-1 capsid. Furthermore, CypA acetylation antagonized the immunosuppressive effects of cyclosporine by inhibiting the sequential steps of cyclosporine binding and calcineurin inhibition. Our results reveal that acetylation regulates key functions of CypA in immunity and viral infection and provide a general set of mechanisms by which acetylation modulates interactions to regulate cell function.

SUBMITTER: Lammers M

PROVIDER: S-EPMC3867001 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND: Cyclophilin A (CypA) represents a potential target for antiretroviral therapy since inhibition of CypA suppresses human immunodeficiency virus type 1 (HIV-1) replication, although the mechanism through which CypA modulates HIV-1 infectivity still remains unclear. The interaction of HIV-1 viral protein R (Vpr) with the human peptidyl prolyl isomerase CypA is known to occur in vitro and in vivo. However, the nature of the interaction of CypA with Pro-35 of N-terminal Vpr has remained undefined. RESULTS: Characterization of the interactions of human CypA with N-terminal peptides of HIV-1 Vpr has been achieved using a combination of nuclear magnetic resonace (NMR) exchange spectroscopy and surface plasmon resonance spectroscopy (SPR). NMR data at atomic resolution indicate prolyl cis/trans isomerisation of the highly conserved proline residues Pro-5, -10, -14 and -35 of Vpr are catalyzed by human CypA and require only very low concentrations of the isomerase relative to that of the peptide substrates. Of the N-terminal peptides of Vpr only those containing Pro-35 bind to CypA in a biosensor assay. SPR studies of specific N-terminal peptides with decreasing numbers of residues revealed that a seven-residue motif centred at Pro-35 consisting of RHFPRIW, which under membrane-like solution conditions comprises the loop region connecting helix 1 and 2 of Vpr and the two terminal residues of helix 1, is sufficient to maintain strong specific binding. CONCLUSIONS: Only N-terminal peptides of Vpr containing Pro-35, which appears to be vital for manifold functions of Vpr, bind to CypA in a biosensor assay. This indicates that Pro-35 is essential for a specific CypA-Vpr binding interaction, in contrast to the general prolyl cis/trans isomerisation observed for all proline residues of Vpr, which only involve transient enzyme-substrate interactions. Previously suggested models depicting CypA as a chaperone that plays a role in HIV-1 virulence are now supported by our data. In detail the SPR data of this interaction were compatible with a two-state binding interaction model that involves a conformational change during binding. This is in accord with the structural changes observed by NMR suggesting CypA catalyzes the prolyl cis/trans interconversion during binding to the RHFP35RIW motif of N-terminal Vpr.

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Acetylation regulates cyclophilin A catalysis, immunosuppression and HIV isomerization.

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