title: Selective blockade of trypanosomatid protein synthesis by a recombinant antibody anti-Trypanosoma cruzi P2β protein.
abstract: The ribosomal P proteins are located on the stalk of the ribosomal large subunit and play a critical role during the elongation step of protein synthesis. The single chain recombinant antibody C5 (scFv C5) directed against the C-terminal region of the Trypanosoma cruzi P2β protein (TcP2β) recognizes the conserved C-terminal end of all T. cruzi ribosomal P proteins. Although this region is highly conserved among different species, surface plasmon resonance analysis showed that the scFv C5 possesses very low affinity for the corresponding mammalian epitope, despite having only one single amino-acid change. Crystallographic analysis, in silico modelization and NMR assays support the analysis, increasing our understanding on the structural basis of epitope specificity. In vitro protein synthesis experiments showed that scFv C5 was able to specifically block translation by T. cruzi and Crithidia fasciculata ribosomes, but virtually had no effect on Rattus norvegicus ribosomes. Therefore, we used the scFv C5 coding sequence to make inducible intrabodies in Trypanosoma brucei. Transgenic parasites showed a strong decrease in their growth rate after induction. These results strengthen the importance of the P protein C terminal regions for ribosomal translation activity and suggest that trypanosomatid ribosomal P proteins could be a possible target for selective therapeutic agents that could be derived from structural analysis of the scFv C5 antibody paratope.
authors.affiliation: Laboratorio de Biología Molecular de la Enfermedad de Chagas, Instituto de Investigaciones en Ingeniería Genética y Biología Molecular, Consejo Nacional de Investigaciones Científicas y Técnicas, Buenos Aires, Argentina.
authors.forename: Maximiliano Juri
authors.initials: MJ
authors.lastname: Ayub
authors.forename: Benson
authors.initials: B
authors.lastname: Nyambega
authors.forename: Leandro
authors.initials: L
authors.lastname: Simonetti
authors.forename: Tomas
authors.initials: T
authors.lastname: Duffy
authors.forename: Silvia A
authors.initials: SA
authors.lastname: Longhi
authors.forename: Karina A
authors.initials: KA
authors.lastname: Gómez
authors.forename: Johan
authors.initials: J
authors.lastname: Hoebeke
authors.forename: Mariano J
authors.initials: MJ
authors.lastname: Levin
authors.forename: Cristian R
authors.initials: CR
authors.lastname: Smulski
mesh_terms: D000913:Antibodies, Protozoan / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology*; D014355:Chagas Disease / Q000188:drug therapy / Q000378:metabolism; D000939:Epitopes / Q000737:chemistry / Q000276:immunology; D015870:Gene Expression; D006801:Humans; D008958:Models, Molecular; D010802:Phylogeny; D011485:Protein Binding / Q000187:drug effects; D014176:Protein Biosynthesis / Q000187:drug effects*; D011487:Protein Conformation; D015800:Protozoan Proteins / Q000096:biosynthesis* / Q000145:classification / Q000235:genetics / Q000276:immunology; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology; D012269:Ribosomal Proteins / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000145:classification / Q000276:immunology; D057127:Single-Chain Antibodies / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology*; D014349:Trypanosoma cruzi / Q000235:genetics / Q000276:immunology / Q000378:metabolism*
chemical_list: D000913:Antibodies, Protozoan; D000939:Epitopes; D015800:Protozoan Proteins; D011994:Recombinant Proteins; D012269:Ribosomal Proteins; D057127:Single-Chain Antibodies