title: Isolation and characterization of gram-positive cyanophycin-degrading bacteria-kinetic studies on cyanophycin depolymerase activity in aerobic bacteria.
abstract: This study is the first report on the extracellular degradation of cyanophycin (CGP) by Gram-positive bacteria. Three different Gram-positive bacteria were isolated from forest soil that were able to utilize CGP as the sole carbon source for growth. The isolates were assigned to species of the genera Bacillus and Micromonospora. From one of the isolates, which was taxonomically affiliated as Bacillus megaterium strain BAC19, the extracellular CGP depolymerase (extracellular CGPase; CphEBm) was purified to electrophoretic homogeneity by fast protein liquid chromatography and affinity binding to an arginine-agarose column. The purified enzyme was specific for hydrolytic cleavage of CGP, and inhibitor studies indicated that CphEBm is a serine-type peptidase. As CGP degradation products, (beta-Asp-Arg)2 tetrapeptides in addition to beta-Asp-Arg dipeptides occurred, which were identified by electrospray ionization mass spectrometry analysis. Furthermore, a novel quantitative enzyme assay was developed for kinetic studies on CGP depolymerases. For CphEBm, as well as for the extracellular CGPase of Pseudomonas anguilliseptica strain BI (CphEPa), KM values of 2.2 and 1.0 microM, respectively, for CGP were determined.
authors.affiliation: Institut für Molekulare Mikrobiologie und Biotechnologie, Westfälische Wilhelms-Universität Münster, Corrensstrasse 3, 48149 Münster, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Obst
authors.forename: Ahmed
authors.initials: A
authors.lastname: Sallam
authors.forename: Heinrich
authors.initials: H
authors.lastname: Luftmann
authors.forename: Alexander
authors.initials: A
authors.lastname: Steinbüchel
mesh_terms: D001407:Bacillus / Q000145:classification / Q000302:isolation & purification / Q000378:metabolism; D001420:Bacteria, Aerobic / Q000145:classification / Q000302:isolation & purification / Q000378:metabolism; D001426:Bacterial Proteins; D001673:Biodegradation, Environmental; D006094:Gram-Positive Bacteria / Q000145:classification / Q000302:isolation & purification / Q000378:metabolism*; D006867:Hydrolases / Q000378:metabolism*; D007700:Kinetics; D008847:Micromonospora / Q000145:classification / Q000302:isolation & purification / Q000378:metabolism; D009842:Oligopeptides / Q000032:analysis; D010940:Plant Proteins / Q000378:metabolism*; D012988:Soil Microbiology
chemical_list: D001426:Bacterial Proteins; D009842:Oligopeptides; D010940:Plant Proteins; C024250:cyanophycin; D006867:Hydrolases