Two novel asparaginyl endopeptidase-like cysteine proteinases from the protist Trichomonas vaginalis: their evolutionary relationship within the clan CD cysteine proteinases
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- 作者：Josefina Leó ; n-Fé ; lix ; Jaime Ortega-Ló ; pez ; Ricardo Orozco-Solı ; ́ ; s ; Rossana Arroyo
- 关键词：Family C13 ; Legumains ; E-64 ; 2-D zymograms ; Aza-peptide-epoxides ; Phylogeny
- 刊名：Gene
- 出版年：2004
- 出版时间：23 June 2004
- 年：2004
- 卷：335
- 期：Complete
- 页码：25-35
- 全文大小：787 K
文摘
Cysteine proteinases (CPs) are important virulence factors of the protozoan parasite Trichomonas vaginalis. A total of six genes coding for cathepsin L-like CPs belonging to clan CA have been identified in T. vaginalis. At least 23 distinct spots with proteolytic activity have been detected by two-dimensional (2-D) substrate gel electrophoresis from in vitro grown parasites; however, only few of them have been characterized. In this work, we detected six spots with proteolytic activity and molecular weights between 25 and 35 kDa. The six proteinases correspond to two distinct CP families: the papain-like family, represented by four spots with pIs between 4.5 and 5.5; and the legumain-like family represented by two spots with pI 6.3 and 6.5. Next, we obtained two cDNAs encoding for legumain-like CPs from T. vaginalis, which were named Tvlegu-1 and Tvlegu-2. The size of these cDNA clones were 1225 and 1364 bp, which encoded for 388 and 415 amino acids, respectively. Their putative translation products have molecular masses of 42.8 and 47.2 kDa, corresponding to inactive legumain-like CP precursors. The two sequences share 40 % identity at the amino acid level. These protein products can be classified within a branch of the legumain-like family in clan CD cysteine proteinases due to their sensitivity to specific proteinases inhibitors, their DNA sequences, and phylogenetic reconstruction. However, they do not correspond either to the typical asparaginyl endopeptidase or the glycosylphosphatidylinositol (GPI): protein transamidase subfamilies. These results suggest that the TVLEGU-1 and TVLEGU-2 peptidases are likely to be part of a new subfamily within the legumain-like family of clan CD cysteine proteinases. Furthermore, they could be one of the missing links between prokaryotic and eukaryotic CPs in clan CD enzymes.