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Crystal Structure of Lyme Disease Variable Surface Antigen VlsE of Borrelia burgdorferi*

Open AccessPublished:March 28, 2002DOI:https://doi.org/10.1074/jbc.M201547200
      VlsE is an outer surface lipoprotein ofBorrelia burgdorferi that undergoes antigenic variation through an elaborate gene conversion mechanism and is thought to play a major role in the immune response to the Lyme disease borellia. The crystal structure of recombinant variant protein VlsE1 at 2.3-Å resolution reveals that the six variable regions form loop structures that constitute most of the membrane distal surface of VlsE, covering the predominantly α-helical, invariant regions of the protein. The surface localization of the variable amino acid segments appears to protect the conserved regions from interaction with antibodies and hence may contribute to immune evasion.
      vls
      variable major protein (VMP)-like sequence
      VR
      variable region
      DR
      direct repeat
      IR
      invariant region
      aa
      amino acids
      Lyme disease is a multistage, tick-borne infection that is endemic to regions of the United States, Europe, and Asia (
      • Steere A.C.
      ). The causative bacteria are a family of closely related spirochetes, includingBorrelia burgdorferi, Borrelia garinii, and Borrelia afzelii, and are transmitted from one mammal to another by Ixodes ticks. Lyme disease borrelia cause persistent infections and chronic neurologic, cardiovascular, and arthralgic manifestations that can last for months to years in humans and other mammals if not treated successfully, indicating that the spirochetes can effectively evade the host's immune defenses.
      The mechanisms of immune evasion are not well understood at this time but are thought to include at least one form of antigenic variation. The variable major protein (VMP)-like sequence (vls)1 locus ofB. burgdorferi (
      • Zhang J.R.
      • Hardham J.M.
      • Barbour A.G.
      • Norris S.J.
      ) is a complex antigenic variation system that in many ways resembles the more thoroughly characterized variable major protein system of relapsing fever borrelia (
      • Barbour A.
      • Restrepo B.
      ). Thevls locus of B. burgdorferi B31 consists of the expression site vlsE and a contiguous set of 15vls silent cassettes. The exact function of the 35-kDa surface-exposed lipoprotein VlsE is unknown; however, it is thought that the vls system may play an important role in mammalian infection because loss of the encoding linear plasmid lp28-1 results in reduced infectivity (
      • Labandeira-Rey M.
      • Baker E.
      • Skare J.T.
      ,
      • Purser J.E.
      • Norris S.J.
      ).
      The overall primary structure of VlsE consists of N-terminal and C-terminal constant domains flanking a central cassette region. The cassette region exhibits ∼90% sequence identity with the silent cassettes, and most of the sequence differences are concentrated in six variable regions (VR1–VR6) interspersed among six invariant regions (IR1–IR6). Both thevlsE cassette region and the silent cassettes are demarcated by 17-bp direct repeats (DR). During experimental infection of mice, segments of the silent cassettes recombine into the vlsEcassette region through a gene conversion mechanism (
      • Zhang J.R.
      • Hardham J.M.
      • Barbour A.G.
      • Norris S.J.
      ,
      • Zhang J.R.
      • Norris S.J.
      ). The sequence changes are detectable within 4 days after experimental infection of mice; by 28 days every isolate from skin or other tissues is unique and contains roughly 9–13 recombination events (
      • Zhang J.R.
      • Norris S.J.
      ). It has been hypothesized that the humoral immune response is unable to respond effectively to the seemingly continuous generation of new VlsE variants, thus permitting immune evasion and persistent infection (
      • Zhang J.R.
      • Hardham J.M.
      • Barbour A.G.
      • Norris S.J.
      ,
      • Zhang J.R.
      • Norris S.J.
      ). Paradoxically, VlsE induces a strong antibody response in Lyme disease patients and infected animals, and recombinant VlsE protein and a peptide corresponding to IR6 have been found to be useful in the serodiagnosis of Lyme disease (
      • Lawrenz M.B.
      • Hardham J.M.
      • Owens R.T.
      • Nowakowski J.
      • Steere A.C.
      • Wormser G.P.
      • Norris S.J.
      ,
      • Liang F.T.
      • Alvarez A.L, Gu, Y.
      • Nowling J.M.
      • Ramamoorthy R.
      • Philipp M.T.
      ,
      • Liang F.T.
      • Steere A.C.
      • Marques A.R.
      • Johnson B.J.
      • Miller J.N.
      • Philipp M.T.
      ). Thus a dilemma has existed between the high immunogenicity of VlsE and its possible role in immune evasion.
      To better understand the relationship between VlsE structure and antigenicity and to gain insight into its possible function(s), we have determined the three-dimensional structure of VlsE from B. burgdorferi B31. For these studies, a recombinant form of VlsE1, the first variant of VlsE identified (
      • Zhang J.R.
      • Hardham J.M.
      • Barbour A.G.
      • Norris S.J.
      ), was utilized. The signal peptide sequence (corresponding to aa 1–19 of the full-length sequence) was omitted from the recombinant construct to permit isolation and crystallization of a protein lacking the lipid moiety. The resulting protein thus contained the N-terminal region (aa 20–115), the cassette region (aa 116–310), and the C-terminal region (aa 311–346) of the mature protein. The recombinant protein also contained an N-terminal polyhistidine sequence to aid in the purification process (
      • Lawrenz M.B.
      • Hardham J.M.
      • Owens R.T.
      • Nowakowski J.
      • Steere A.C.
      • Wormser G.P.
      • Norris S.J.
      ).

      ACKNOWLEDGEMENTS

      We thank the Advanced Photon Source beamline operators for assistance at beamline 14-BM-D and Julie C. Holding for excellent assistance in the laboratory.

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