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      A Novel MHC-I Surface Targeted for Binding by the MCMV m06 Immunoevasin Revealed by Solution NMR.

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          Abstract

          As part of its strategy to evade detection by the host immune system, murine cytomegalovirus (MCMV) encodes three proteins that modulate cell surface expression of major histocompatibility complex class I (MHC-I) molecules: the MHC-I homolog m152/gp40 as well as the m02-m16 family members m04/gp34 and m06/gp48. Previous studies of the m04 protein revealed a divergent Ig-like fold that is unique to immunoevasins of the m02-m16 family. Here, we engineer and characterize recombinant m06 and investigate its interactions with full-length and truncated forms of the MHC-I molecule H2-L(d) by several techniques. Furthermore, we employ solution NMR to map the interaction footprint of the m06 protein on MHC-I, taking advantage of a truncated H2-L(d), "mini-H2-L(d)," consisting of only the α1α2 platform domain. Mini-H2-L(d) refolded in vitro with a high affinity peptide yields a molecule that shows outstanding NMR spectral features, permitting complete backbone assignments. These NMR-based studies reveal that m06 binds tightly to a discrete site located under the peptide-binding platform that partially overlaps with the β2-microglobulin interface on the MHC-I heavy chain, consistent with in vitro binding experiments showing significantly reduced complex formation between m06 and β2-microglobulin-associated MHC-I. Moreover, we carry out NMR relaxation experiments to characterize the picosecond-nanosecond dynamics of the free mini-H2-L(d) MHC-I molecule, revealing that the site of interaction is highly ordered. This study provides insight into the mechanism of the interaction of m06 with MHC-I, suggesting a structural manipulation of the target MHC-I molecule at an early stage of the peptide-loading pathway.

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          Author and article information

          Journal
          J. Biol. Chem.
          The Journal of biological chemistry
          American Society for Biochemistry & Molecular Biology (ASBMB)
          1083-351X
          0021-9258
          Nov 27 2015
          : 290
          : 48
          Affiliations
          [1 ] From the Laboratory of Chemical Physics, NIDDK, and.
          [2 ] the Molecular Biology Section, Laboratory of Immunology, NIAID, National Institutes of Health, Bethesda, Maryland 20892.
          [3 ] From the Laboratory of Chemical Physics, NIDDK, and bax@nih.gov.
          [4 ] the Molecular Biology Section, Laboratory of Immunology, NIAID, National Institutes of Health, Bethesda, Maryland 20892 dhm@nih.gov.
          Article
          M115.689661
          10.1074/jbc.M115.689661
          4661401
          26463211
          e926e32e-649c-4af4-a1fa-d184ac48902d
          History

          MCMV,antigen presentation,herpesvirus,immunoevasin,m06/gp48,major histocompatibility complex (MHC),nuclear magnetic resonance (NMR),protein-protein interaction,surface plasmon resonance (SPR)

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