Degradation of chitosans with a family 46 chitosanase from Streptomyces coelicolor A3(2).
We have studied the degradation of well-characterized soluble heteropolymeric chitosans by a novel family 46 chitosanase, ScCsn46A from Streptomyces coelicolor A3(2), to obtain insight into the enzyme's mode of action and to determine its potential for production of different chitooligosaccharides. The degradation of both a fully deacetylated chitosan and a 32% acetylated chitosan showed a continuum of oligomeric products and a rapid disappearance of the polymeric fraction, which is diagnostic for a nonprocessive endomode of action. The kinetics of the degradation of the 32% acetylated chitosan demonstrated an initial rapid phase and a slower second phase, in addition to a third and even slower kinetic phase. The first phase reflects the cleavage of the glycosidic linkage between two deacetylated units (D-D), the primary products being fully deacetylated dimers, trimers, and tetramers, as well as longer oligomers with increasing degrees of acetylation. In the subsequent slower kinetic phases, oligomers with a higher degree of acetylated units (A) appear, including oligomers with A's at the reducing or nonreducing end, which indicate that there are no absolute preferences for D in subsites -1 and +1. After maximum degradation of the chitosan, the dimers DA and DD were the dominant products. The degradation of chitosans with varying degrees of acetylation to a maximum degree of scission showed that ScCsn46A could degrade all chitosan substrates extensively, although to decreasing degrees of scission with increasing F(A). The potential use of ScCsn46A to prepare fully deacetylated oligomers and more highly acetylated oligomers from chitosan substrates with varying degrees of acetylation is discussed.
Heggset
Ellinor B
EB
Norwegian Biopolymer Laboratory (NOBIPOL), Department of Biotechnology, Norwegian University of Science and Technology, 7491 Trondheim, Norway.
Dybvik
Anette I
AI
Hoell
Ingunn A
IA
Norberg
Anne Line
AL
Sørlie
Morten
M
Eijsink
Vincent G H
VG
Vårum
Kjell M
KM
D000107:Acetylation; D001672:Biocompatible Materials / Q000737:chemistry / Q000378:metabolism*; D002688:Chitinases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D048271:Chitosan / Q000737:chemistry / Q000378:metabolism*; D002850:Chromatography, Gel; D004269:DNA, Bacterial / Q000235:genetics; D009682:Magnetic Resonance Spectroscopy; D009844:Oligosaccharides / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D048372:Streptomyces coelicolor / Q000201:enzymology*
D001672:Biocompatible Materials; D004269:DNA, Bacterial; D009844:Oligosaccharides; D011994:Recombinant Proteins; D048271:Chitosan; D002688:Chitinases