Caracterização estrutural da proteína Sm21.7 recombinante de Schistosoma mansoni

• Ano de defesa: 2014
• Autor(a) principal: Mariana Amalia Figueiredo Costa
• Tipo de documento: Tese
• Tipo de acesso: Acesso aberto
• Idioma: por
• Instituição de defesa: Universidade Federal de Minas Gerais; UFMG
• Palavras-chave em Português: Bioquímica e Imunologia; Bioquímica
• Link de acesso: http://hdl.handle.net/1843/BUBD-A2FGCX

Schistosomiasis is an inflammatory chronic disease that represents a major health problem in tropical and subtropical countries. The drug of choice for the treatment, Praziquantel, is effective in killing adult worms, but fails to kill immature forms and prevent reinfection. One important candidate to compose an antischistosomiasis vaccine is the protein Sm21.7 from Schistosoma mansoni, that is capable to reduce the worm burden in a murine immunization model. Sm21.7 has an Nterminal EF hand domain and a C-terminal dynein light chain domain. In the present work, the Sm21.7 gene was cloned and the recombinant protein (rSm21.7) expressed in Escherichia coli and purified to homogeneity. Circular Dichroism analysis showed that the protein was partly folded with 37% of -helices and 15% of -strands. rSm21.7 showed to behave as a dimer in a cross-linking with glutaraldehyde experiment. Crystals of rSm21.7 suitable for X-ray diffraction studies were obtained using PEG monomethyl ether 2,000 and PEG 8,000 as precipitants in diferent conditions. X-ray diffraction images of a native crystal (at 2.05 Å resolution) and a NaI derivative (at 1.95 Å resolution) were collected at W01B-MX2 beamline at the Laboratório Nacional de Luz Síncrotron (LNLS). Both crystals belong to the hexagonal system with similar unit cell parameters a=b=108.5, c= 55.8 Å. SIRAS-derived phases were used to generate the first electron density map where a partial model of rSm21.7 was automatically constructed. The model consists of 96 C-terminal amino acid residues of the full-length protein (from Gln89 to Asn184 dynein light chain domain) and the cleavage of the protein in the crystallization drop was confirmed by SDS-PAGE using sample from dissolved crystals. The Dynein Light Chain domain which was crystallized has 2 - helices and 4 -strands that form an antiparallel -sheet. The structure of the rSm21.7 C-terminal domain allowed the localization of epitopes responsible for the protective response elicited by the protein.