The Diagnostic Performance of Recombinant Trypanosoma cruzi Ribosomal P2b Protein is Influenced by its Expression System

MARCIPAR, I. S.; OLIVARES, M. L.; ROBLES, L.; DEKANTY, A. M.; SILBER, A. M.

PROTEIN EXPRESSION AND PURIFICATION

Elsevier

Año: 2004 vol. 34 p. 1 - 7

1046-5928

In the present work, we have determined the effect of expression vectors and their corresponding host bacteria on the antigenic performance of Trypanosoma cruzi P2b (TcP2b) full-length recombinant protein. The gene encoding the TcP2b ribosomal protein was cloned in pMAL-c2 and pET-32a vectors that allow the expression of high levels of soluble fusion proteins. A panel of 32 negative sera was assayed with the purified proteins expressed using pMal-c2 (TcP2b-MBP) and pET-32a (TcP2b-TRX) vectors and with MBP and TRX purified from pMAL-c2 and pET-32a vectors, respectively. The antigenic behavior of  each Tcp2b recombinant protein differed in the diagnostic performance in terms of DI(+) (93.7 for TcP2b-MBP vs 100% for TcP2b-TRX), in DI(-) (90.5 TcP2b-MBP vs 100% for TcP2b-TRX) and in cross-reaction with negative sera. To determine if the higher reactivity of expressed pMAL-c2 protein was due to folding during  protein expression or to steric effect related to the protein adsorption at the titration plate, the reactivity of sera against soluble proteins was assessed by ELISA inhibition assays. As each soluble protein preserved its level of reactivity, we concluded that differences in reactivity were due to intrinsic characteristics of the proteins and not to differences in patterns of adsorption to the plates.