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Initiation of prolyl cis-trans isomerisation in the CDR-H3 loop of an antibody in response to antigen binding.

ABSTRACT: Proline cis-trans isomerisation is a regulatory mechanism used in a range of biological processes, and is related to various diseases such as Alzheimers disease and cancer. However, the details of the exact molecular mechanism by which it occurs are not known. Using X-ray crystallography, proline isomerisation has been shown to occur following formation of an antigen-antibody complex between the target epiregulin (EPR) and the antibody 9E5, at proline (Pro103), located in the third complementarity-determining region (CDR) of the heavy chain of 9E5. To obtain an accurate description of the pathway involved in cis-trans isomerisation in this system, we performed ten independent long molecular dynamics (MD) simulations starting at a stable transient bound structure obtained from many short binding MD simulations. As a result, we were able to describe the process by which cis-trans isomerisation is initiated, and suggest a catalysis mechanism for cis-trans isomerization in this antigen-antibody system. We found that Asp102, which is immediately adjacent to Pro103, rotates while changing its interacting partner residues in the light chain of 9E5, and at the same time EPR polar residues help to stabilise the intermediate states in the isomerisation process by interacting strongly with Asp102.

SUBMITTER: Shinoda K

PROVIDER: S-EPMC5717248 | biostudies-other | 2017 Dec

REPOSITORIES: biostudies-other

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Initiation of prolyl cis-trans isomerisation in the CDR-H3 loop of an antibody in response to antigen binding.

Shinoda Keiko K Fujitani Hideaki H

Scientific reports 20171205 1

Proline cis-trans isomerisation is a regulatory mechanism used in a range of biological processes, and is related to various diseases such as Alzheimers disease and cancer. However, the details of the exact molecular mechanism by which it occurs are not known. Using X-ray crystallography, proline isomerisation has been shown to occur following formation of an antigen-antibody complex between the target epiregulin (EPR) and the antibody 9E5, at proline (Pro103), located in the third complementari ...[more]

PMID: 29208911

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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.

Dataset Information

Initiation of prolyl cis-trans isomerisation in the CDR-H3 loop of an antibody in response to antigen binding.

Publications

Initiation of prolyl cis-trans isomerisation in the CDR-H3 loop of an antibody in response to antigen binding.

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