STRUCTURAL CHARACTERIZATION OF THE ANAPLASMA MARGINALE ANTIGENICALLY VARIANT MSP2 PROTEIN
Graça, Telmo Gonçalo Henriques
MetadataShow full item record
Antigenic variation allows pathogens to escape host immune responses and establish persistent infection. Anaplasma marginale uses gene conversion from a genomic allelic repertoire to generate msp2 expression site variants. Expressed variants are predominantly derived from whole donor alleles during early infection; in contrast, the majority of variants expressed during persistent infection are mosaics, derived by recombination of oligonucleotide segments from multiple alleles. Prior research had identified that variation occurred within a surface-exposed hypervariable domain (HVR) and specifically within immunogenic HVR microdomains. My research addressed two major knowledge gaps related to generation of Msp2 HVR microdomain variants. The first was to test whether single segmental changes in the HVR microdomains are sufficient to evade a broad antibody response. Collectively, 89% of the segmental variants were true antigenic variants with a single amino acid substitution being sufficient to generate a variant. Biological relevance is supported by the following: (i) all structural variants were expressed during infection of a natural host, (ii) the structural variation observed in the microdomains corresponded to the mean length of variants generated by segmental gene conversion, and (iii) antigenic variants were identified using a broad antibody response. The second knowledge gap was to determine the structural basis that allowed recombination of microdomains derived from multiple alleles to assemble a functional outer membrane protein. Using an integrated approach of predictive modeling with determination of native Msp2 protein structure and function, I established that structured elements, most notably β-sheets, are significantly concentrated in the highly conserved N- and C-terminal domains. In contrast the HVR is overwhelmingly random coil with the structured α-helices and β-sheets confined to the genomically defined “structural tethers” that separate the antigenically variable microdomains. This structure is supported by the surface exposure of the HVR microdomains and the slow diffusion type porin function in native Msp2. In summary, my research demonstrated that segmental gene conversion efficiently creates true Msp2 antigenic variants and that the HVR structure provides plasticity for formation of functional HVR mosaics and realization of the full potential of segmental gene conversion to dramatically expand the variant repertoire.