Mode of action of RG-hydrolase and RG-lyase toward rhamnogalacturonan oligomers. Characterization of degradation products using RG-rhamnohydrolase and RG-galacturonohydrolase.
The mode of action of RG-hydrolase and RG-lyase toward purified linear rhamnogalacturonan (RG) oligomers has been studied. Major tools in the characterization of the degradation products were the exo-acting RG-rhamnohydrolase and RG-galacturonohydrolase. They were used to prepare a series of standards of RG oligomers for HPAEC. 1H NMR spectroscopy confirmed the structure assignment made using HPAEC for a selection of isolated degradation products. Identification of degradation products from purified RG oligomers was then performed by comparing retention times of HPAEC peaks with those of standards. RG-hydrolase was able to cleave RG oligomers which contained five Rha units or more, i.e. DP 9 with a Rha unit at both nonreducing and reducing end. Its preferential cleavage site was at four units from the first nonreducing Rha. RG-lyase was active toward oligomers that contained at least six GalA units, i.e. DP 12 with a GalA at the nonreducing and a Rha at the reducing end. The preferential cleavage site was for the smaller oligomers four residues, and for the largest oligomer six residues from the reducing Rha. From the observed cleavage patterns it can be speculated that in hairy regions, the RG stretches have to be at least 13 residues long for RG-hydrolase and 16 residues long for RG-lyase in order to produce one tetramer.
Mutter
M
M
Wageningen Agricultural University, Department of Food Science, The Netherlands.
Renard
C M
CM
Beldman
G
G
Schols
H A
HA
Voragen
A G
AG
D002851:Chromatography, High Pressure Liquid; D006026:Glycoside Hydrolases / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D008956:Models, Chemical; D010368:Pectins / Q000378:metabolism*; D011108:Polymers / Q000378:metabolism; D011133:Polysaccharide-Lyases / Q000378:metabolism*
D011108:Polymers; D010368:Pectins; D006026:Glycoside Hydrolases; C090358:rhamnogalacturonan rhamnohydrolase; D011133:Polysaccharide-Lyases; C098240:rhamnogalacturonase B