Leishmania donovani: Expression and Characterization of Escherichia coli-Expressed Recombinant Chitinase LdCHT1

• Published in: Experimental parasitology 2001-12, Vol.99 (4), p.220-225
• Main Authors: Razek-Desouky, Abdel, Specht, Charles A., Soong, Lynn, Vinetz, Joseph M.
• Format: Article
• Language: English
• Subjects: Acetylglucosamine - analogs & derivatives; Acetylglucosamine - metabolism; Animals; Biological and medical sciences; Chitin - metabolism; Chitinases - genetics; Chitinases - isolation & purification; Chitinases - metabolism; Chromatography, Gel; Chromatography, Thin Layer; Escherichia coli; Escherichia coli - enzymology; Escherichia coli - genetics; Fluorometry; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation, Enzymologic; Hydrogen-Ion Concentration; Hydrolysis; Hymecromone - analogs & derivatives; Hymecromone - metabolism; Leishmania donovani; Leishmania donovani - enzymology; Leishmania donovani - genetics; Life cycle. Host-agent relationship. Pathogenesis; parasite-vector interaction; promastigote; Protozoa; Psychodidae; recombinant protein expression; Recombinant Proteins - genetics; Recombinant Proteins - isolation & purification; Recombinant Proteins - metabolism; sandfly; Substrate Specificity
• ISSN: 0014-4894; 1090-2449
• DOI: 10.1006/expr.2001.4665

Razek-Desouky, A., Specht, C. A., Soong, L., and Vinetz, J. M. 2001. Leishmania donovani: Expression and characterization of Escherichia coli-expressed recombinant chitinase LdCHT1. Experimental Parasitology99, 220–225. Leishmania parasites produce chitinase activity (EC. 3.2.1.14) thought to be important in parasite-sandfly interactions and transmission of the parasite to the vertebrate host. Previous observations have suggested that parasite chitinases are involved in degradation of the sandfly peritrophic matrix and the chitinous layer of the cardiac valve cuticle. This chitinase activity is thought to produce an incompetent pharyngeal valve sphincter and a route of egress that allow Leishmania promastigotes to be regurgitated into the site of blood feeding. In the studies reported here, enzymatically active L. donovani chitinase LdCHT1 was expressed as a thioredoxin fusion protein in Escherichia coli strain AD494 (DE3). Recombinant LdCHT1 had a predominantly endochitinase activity, in contrast to previous reports of both exo- and endochitinase activities in axenic culture supernatants of diverse Leishmania spp. promastigotes. The predominant endochitinase activity of recombinant LdCHT1 is consistent with the presumed function of the enzyme in disrupting chitinous structures in the sandfly digestive system to allow transmission.