G15SecL05Tpc3: Difference between revisions

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Michael Pape, Farbod Raegan, Michal Harel

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 Crytsallization, binding and X-ray diffraction analysis of Osp-B along with a sampling of its complement-independent antibodies have shown that its structure is analogous to that of Osp-A, with a significant difference: within each borrelia species, Osp-A is largely invariant in amino acid sequence and antigenic reactivity, but Osp-B varies significantly (Becker, 2005). These variations inhibit the ability of scientists to develop Osp-B as a protective vaccine against Lyme borreliosis. Osp-B’s variations are primarily expressed as significant differences of reactivity with monoclonal antibodies and truncation of the C-terminus; this proves problematic for vaccine development since most protective antibodies targeted against Osp-A and Osp-B are generally directed toward this terminus region. Variations such as truncation in Osp-B can inhibit this process making the vaccine ineffective, though certain complement independent antibodies such as the samples examined in the crystallization process have been found to bind and lyse Osp-B in the absence of complement.
 H6831 is a specific IgG class antigen binding fragment that has a series of significant residues that bind to the epitope on Osp-B and lyse it without the aid of a complement, resulting in a unique molecular interaction that causes a physical change in the shape of Osp-B upon binding. In order to better analyze the changes, crystallized structures of Osp-B and the Osp-B H6831 complex were taken and comparatively analyzed: X-ray diffraction showed that the structure of OspB-CT ('''GREEN LINK THAT''') contains twelve anti parallel beta strands and a single alpha helix (Becker, 2005). Strands 1-4 form the Osp-B’s central sheet: this is a free standing sheet that is located at the N-terminus of the protein. The remaining strands 5-12 form two additional sheets that together with the alpha helix form a barrel-like domain. Analysis of the bound complex revealed that the central beta sheet is either destroyed or removed by proteolysis upon binding (Becker, 2005). Ultimately the structural changes to Osp-B appear to result only indirectly from the addition of Fab (Becker, 2005).
-Almost the entirety of this binding process is centered around distinct residues that were discovered due to comparative analysis of Osp-A and Osp-B’s crystal structures. A charged triad is formed in both structures whose united physical form presents itself as a favorable binding site. In Osp-B, this triad consists of amino-acid residues Arg-162, Glu-184, and Arg-214. These triads form a prominent cleft with density void space in between each other. This space is well suited for an all trans-structures to bind into, and such a structure was located on the crystallized Osp-B-H6831 bound complex.  This complex is a combination of Osp-B-CT residues 202-296 and the Fabs’ heavy and light chains. A notable feature of the OspB-CT-H6831 interaction is the prevalence of aromatic residues contributed by the Fab: these aromatic residues are the points at which Lys-253 on Osp-B guides itself to bind with Glu-50. Specifically, Lys 253 wedges itself between a tyrosine and a tryptophan to be able reach the Glu-50 so the binding between the two residues can occur (Becker, 2005). This interaction, along with an amine hydrogen bond and ion pair with a glutamate from the Fab heavy chain, a second hydrogen bond with a main chain carbonyl from loop 1 of Osp-B-CT, and a water mediated hydrogen bond to a histidine from the Fab heavy chain constitute the three main series of interactions in the OspB-H6831 bound complex. Lys-253 is the absolute key component of the formation of this bound complex; if Lys-253 were to be substituted with another amino acid or be missing entirely via external influential mutation, the binding could not occur***.  Multiple studies highlight selective pressures related to this residue: several mutated ''B. burgdorferi'' spirochetes have been shown to lack the Lys-253 residue entirely, though this form of the disease is significantly less pathogenic as a consequence.
+Almost the entirety of this binding process is centered around distinct residues that were discovered due to comparative analysis of Osp-A and Osp-B’s crystal structures. A charged triad is formed in both structures whose united physical form presents itself as a favorable binding site. In Osp-B, this triad consists of amino-acid residues Arg-162, Glu-184, and Arg-214. These triads form a prominent cleft with density void space in between each other. This space is well suited for an all trans-structures to bind into, and such a structure was located on the crystallized Osp-B-H6831 bound complex.  This complex is a combination of Osp-B-CT residues 202-296 and the Fabs’ heavy and light chains. A notable feature of the OspB-CT-H6831 interaction is the prevalence of aromatic residues contributed by the Fab: these aromatic residues are the points at which Lys-253 on Osp-B guides itself to bind with Glu-50. Specifically, Lys 253 wedges itself between residues Try-33 and Tyr-101 of the H6831 heavy chain to be able reach the Glu-50 so the ion-pair binding between the two residues can occur (Becker, 2005). This interaction, along with an amine hydrogen bond and ion pair with a glutamate from the Fab heavy chain, a second hydrogen bond with a main chain carbonyl from loop 1 of Osp-B-CT, and a water mediated hydrogen bond to a histidine from the Fab heavy chain constitute the three main series of interactions in the OspB-H6831 bound complex. Lys-253 is the absolute key component of the formation of this bound complex; if Lys-253 were to be substituted with another amino acid or be missing entirely via external influential mutation, the binding could not occur***.  Multiple studies highlight selective pressures related to this residue: several mutated ''B. burgdorferi'' spirochetes have been shown to lack the Lys-253 residue entirely, though this form of the disease is significantly less pathogenic as a consequence.
 Further studies into this protein and complement-independent antibodies are important since as of now vaccinations of Osp-B have proven unreliable due to regional variations in its form. Creation of a proper vaccine has gradually grown in interest because the current antibodies created in the human body from Osp-A have been shown to have possible correlation with arthritis, an inflammatory joint disorder. In this regard, bactericidal antibodies to Osp-B may have in vivo relevance in late stage diseases and in vaccines for the elimination of spirochetes from ticks feeding on immune individuals (Escudero, 1997).