Partially esterified oligogalacturonides are the preferred substrates for pectin methylesterase of Aspergillus niger.
Investigations on the mode of action of Aspergillus niger pectin methylesterase (PME) towards differently C(6)- and C(1)-substituted oligogalacturonides (oligoGal p A) are described. De-esterification of methyl-esterified (un)saturated oligoGal p A proceeds via a specific pattern, depending on the degree of polymerization. Initially, a first methyl ester of the oligomer is hydrolysed, resulting in one free carboxyl group. Subsequently, this first product is preferred as a substrate and is de-esterified for a second time. This product is then accumulated and hereafter de-esterified further to the final product, i.e. oligoGal p A containing one methyl ester located at the non-reducing end residue for both saturated and unsaturated oligoGal p A, as found by post-source decay matrix-assisted laser-desorption/ionization-time-of-flight MS. The saturated hexamer is an exception to this: three methyl esters are removed very rapidly, instead of two methyl esters. When unsaturated oligoGal p A were used, the formation of the end product differed slightly, suggesting that the unsaturated bond at the non-reducing end influences the de-esterification process. In vivo, PME prefers methyl esters, but the enzyme appeared to be tolerant for other C(6)- and C(1)-substituents. Changing the type of ester (ethyl esterification) or addition of a methyl glycoside (C(1)) only reduced the activity or had no effect respectively. The specific product pattern was identical for all methyl- and ethyl-esterified oligoGal p A and methyl-glycosidated oligoGal p A, which strongly indicates that one or perhaps two non-esterified oligoGal p A are preferred in the active-site cleft.
van Alebeek
Gert-Jan W M
GJ
Laboratory of Food Chemistry, Department of Agrotechnology and Food Sciences, Wageningen University, Bomenweg 2, 6703 HD Wageningen, The Netherlands.
van Scherpenzeel
Katrien
K
Beldman
Gerrit
G
Schols
Henk A
HA
Voragen
Alphons G J
AG
D001234:Aspergillus niger / Q000201:enzymology*; D002265:Carboxylic Ester Hydrolases / Q000378:metabolism*; D004952:Esters; D009844:Oligosaccharides / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013379:Substrate Specificity
D004952:Esters; D009844:Oligosaccharides; C412763:oligogalacturonic acid; D002265:Carboxylic Ester Hydrolases; C022323:pectinesterase