title: Deficiency of current methods in assaying endochitinase activity.
abstract: Since chitin is degraded by a combination of both endo- and exochitinases, it is likely that both enzymes will be present in a crude extract. Currently used substrates for detecting endochitinase activity suffer from the fact that they could easily be digested by the contaminating exochitinase, thus giving either a false-positive or an inaccurate reading of the endochitinase activity. Using Photorhabdus luminescens, a bacterium symbiotically associated with insect-parasitic nematode Heterorhabditis bacteriophora as an exemplary system, we have identified these two chitinases by a simple "fluorimetric zymography" procedure. The exochitinase is a metalloenzyme and its activity is inhibited by 1,10-phenanthroline. Once the exochitinase activity is detected in a crude extract, its contribution must be eliminated before accurate determination of the endochitinase activity can be carried out. Specific properties of these enzymes including the pH activity profile, the requirement of metal ions for activity, and the molecular weight of the enzymes are among the factors to be considered in developing assaying procedures for endochitinase.
authors.affiliation: Department of Biology, Temple University, Philadelphia, Pennsylvania, 19122, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Ren
authors.forename: K E
authors.initials: KE
authors.lastname: Wee
authors.forename: F N
authors.initials: FN
authors.lastname: Chang
mesh_terms: D002688:Chitinases / Q000032:analysis*; D002845:Chromatography; D005470:Fluorometry / Q000379:methods; D006596:Hexosaminidases / Q000032:analysis*; D006863:Hydrogen-Ion Concentration; D020637:Photorhabdus / Q000201:enzymology*
chemical_list: C079639:1,4-beta-chitobiosidase; D006596:Hexosaminidases; D002688:Chitinases