Cathepsin L-like cysteine proteases from Brugia malayi: cDNA cloning and comparison with Caenorhabditis elegans

Abstract

Proteases are thought to be potentially useful targets for developing medicines to control fi lariasis caused by parasitic nematodes. To screen cysteine proteases essential for viability of nematodes, fi fteen cathepsin B/L-like genes of Caenorhabditis elegans, as a model of the parasitic nematodes, were interfered by RNAi. As a result, ~100% embryonic lethality was observed only when Cecpl-1, encoding a cathepsin L-like protease, was knocked down. Subsequent attempts were made to identify the orthologs of Ce-cpl-1 in the parasitic nematode Brugia malayi by molecular cloning and sequencing of some EST clones. The sequences of fi ve distinct open reading frames were identifi ed, all of which are most homologous to Ce-CPL-1 in C. elegans. The consensus catalytic triad of cathepsin L-like proteases is conserved among four of the fi ve predicted proteins. Phylogenetic analysis suggests that one of them, Bm-CPL-1, is closely related to the proteases from other fi larial parasites. However, neither Bm-CPL-1 nor the other four predicted proteins belong to the same sub-branch of Ce-CPL-1 in the phylogenetic tree. In B. malayi, the functions of Ce-CPL-1 are presumably shared by some of the predicted proteases including Bm-CPL-1, although the possibility cannot be ruled out that B. malayi has an unknown cysteine protease more resembling Ce-CPL-1.Lymphatic fi lariasis, caused by the nematode Brugia malayi and Wuchereria bancrofti, is one of the serious public health problems in the tropical countries and billions of people are exposed to the risk of infection worldwide (25, 37). These parasites depend on proteolysis for their life cycle including tissue penetration and evasion from host immune responses (35). Thus some of the proteases are excellent potential targets for the design of novel anti-parasitic drugs. Aspartic proteases of peptidase family A1 (the classifi cation is according to Barrett et al. (3)) were found to be useful as the key therapeutic targets for malaria, schistosomasis etc. (7, 13), and we previously performed molecular cloning and analysis of tissue distribution of those enzymes from B. malayi as the fi rst step toward this direction (2). Besides, cysteine proteases of peptidase family C1 are

Molecular identification and functional characterization of the cathepsin B gene (Ab-cb-1) in the plant parasitic nematode Aphelenchoides besseyi

“…The triad of Cys, His, and Asn residues constitutes an active catalytic domain, which is highly functional and conserved among the cysteine proteases of peptidase family C1, including cathepsin B [ 20 ]. The occluding loop is characteristic of only cathepsin B; this loop is responsible for endopeptidase and exopeptidase activity, and its presence seems to indicate that cathepsin B possesses the ability to act [ 21 , 22 , 23 ] as a self-inhibitor by partially blocking the active site when the loop is inactive [ 24 – 28 ]. These structural features indicated that Ab- CB-1 had the functions and activities of cathepsin B. Ab- CB-1 was inferred to react with substrates directly without protein transportation after synthesis in cells because it did not contain any signal peptide sequence and because the transmembrane signal was weak in the putative transmembrane region.…”