Performance of selected microbial pectinases on synthetic monomethyl-esterified di- and trigalacturonates

Two monomethyl esters of alpha-(1-4)-linked D-galacturonic dimers and three monomethyl esters of alpha-(1-4)-linked D-galacturonic acid trimers were s ynthesized chemically and further used as substrates in order to establish the substrate specificity of six different endopolygalacturonases from Aspe rgillus niger, one exopolygalacturonase from Aspergillus tubingensis, and f our selected Erwinia chrysanthemi pectinases; exopolygalacturonan hydrolase X (PehX), exopolygalacturonate lyase X (PelX), exopectate lyase W (PelW), and oligogalacturonan lyase (Ogl). All A. niger endopolygalacturonases (PGs ) were unable to hydrolyze the two monomethyldigalacturonates and 2-methylt rigalacturonate, whereas 1-methyltrigalacturonate was only cleaved by PGI, PGII, and PGB albeit at an extremely low rate. The hydrolysis of 3-methyltr igalacturonate into 2-methyldigalaeturonate and galacturonate by all endopo lygalacturonases demonstrates that these enzymes can accommodate a methylga lacturonate at subsite -2. The A. tubingensis exopolygalacturonase hydrolyz ed the monomethyl-esterified digalacturonates and trigalacturonates althoug h at lower rates than for the corresponding oligogalacturonates. 1-Methyltr igalacturonate was hydrolyzed at the same rate as trigalacturonate which de monstrates that the presence of a methyl ester at the third galacturonic ac id from the nonreducing end does not have any effect on the performance of exopolygalacturonase. Of the four E. chrysanthemi pectinases, Ogl was the o nly enzyme able to cleave digalacturonate, whereas all four enzymes cleaved trigalacturonate. Ogl does not cleave monomethyl-esterified digalacturonat e and trigalacturonate in case the second galacturonic acid residue from th e reducing end is methyl-esterified. PehX did not hydrolyze any of the mono methyl-esterified trigalacturonates. The two lyases, PelX and PelW, were bo th only able to cleave 1-methyltrigalacturonate into Delta 4,5-unsaturated 1-methyldigalacturonate and galacturonate.