Oligopeptidase B from Leishmania amazonensis: molecular cloning, gene expression analysis and molecular model

Abstract

Serine oligopeptidases of trypanosomatids are emerging as important virulence factors and therapeutic targets in trypanosome infections. A complete open reading frame of oligopeptidase B from Leishmania amazonensis was amplified with polymerase chain reaction with gradient annealing temperatures using primers designed for the oligopeptidase B gene from L. major. The 2,196-bp fragment coded for a protein of 731 amino acids with a predicted molecular mass of 83.49 KDa. The encoded protein (La_OpB) shares a 90% identity with oligopeptidases of L. major and L. infantum, 84% with L. braziliensis, and approximately 62% identity with Trypanosoma peptidases. The oligopeptidase B gene is expressed in all cycle stages of L. amazonensis. The three dimensional model of La_OpB was obtained by homology modeling based on the structure of prolyl oligopeptidases. We mapped a La_OpB model that presents a greater negative charge than prolyl oligopeptidases; our results suggest a difference in the S2 subsite when compared to oligopeptidases B from Trypanosoma and bacterial oligopeptidases B. The La_OpB model serves as a starting point for its exploration as a potential target source for a rational chemotherapy.

Oligopeptidase B-2 from Leishmania amazonensis with an unusual C-terminal extension

“…As observed in T. cruzi, the results showed that the oligopetidase B-2 gene of L. amazonensis is expressed in both procyclic and metacyclic promastigotes and in axenic amastigotes suggesting that the gene is constitutively expressed throughout the life cycle of the parasite. The same results were observed with the oligopeptidase B gene of L. amazonensis (Guedes et al 2007) and with ITS region used as control (not shown).…”

“…(75%) is less than observed between oligopeptidases B-2 of T. cruzi and T. brucei (82%). Interestingly, oligopeptidases B-2 is more conserved than classical trypanosomatid oligopeptidases B (Guedes et al 2007). Sequence alignments with oligopeptidases B-2 from other trypanosomatids indicated conservation of the entire sequence with an absence of gaps (not shown).…”

“…The encoded oligopeptidase B-2 from L. amazonensis shared ~21% identity with bacterial oligopeptidase B (Table I), while oligopeptidase B from L. amazonensis shared 36% identity (Guedes et al 2007).…”

Serine Proteases and Vaccines against Leishmaniasis: A Dual Role

“…From the sequencing of the L. major genome, a number of distinct serine proteases have been identified: (i) subtilisin-like, an S9 family member that also includes prolyloligopeptidase (POP), peptidyl-1 dipeptidase IV and oligopeptidase B (OPB), (ii) a type I signal peptidase (SPase I), (iii) a lysosomal serine carboxypeptidase, (iv) a 26S regulatory proteasome subunit, (v) a nucleoporin homolog and (vi) an orthologous of the rhomboid-like intramembranous serine peptidase family [12]. From the L. amazonensis genome, two others of interest are; (vii) Oligopeptidase B [13] and (viii) Oligopeptidase B2 [14].…”

“…Oligopetidase B, along with a new member called have been identified in L. amazonensis [13,14], however, their biological roles are unknown. Attempts to create null mutants of SPase I have been unsuccessful, suggesting that this enzyme is crucial to survival.…”