Model for β-1,6-N-acetylglucosamine oligomer hydrolysis catalysed by DispersinB, a biofilm degrading enzyme

DispersinB (DspB), a member of beta-1,6-N-acetylglucosaminidase group of GH 20 glycoside hydrolases, catalyses the biofilm degradation of several human pathogenic microorganisms. DspB is a (beta/alpha)(8) barrel protein, showing retaining cleavage mechanism towards oligomer and polymer substrates. A chromophore containing oligomer substrate series was used to study the DspB's mode of action. The hydrolysis reaction of beta(1,6)-linked N-acetylglucosamine thiophenyl glycosides with degree of polymerisation of 2, 3, 4 and 5 was followed by reversed phase HPLC and progress curves were determined and analysed. Based on the analysis of process curves obtained from prolonged hydrolysis we assumed the presence of more productive binding modes resulting in parallel reactions followed by consecutive reaction steps. Strictly nonreducing-end specificity was observed, the presence of monomer, dimer and trimer non-reducing-end products was verified by MALDI-TOF MS. Another cleavage was suggested after the first glycosidic attack in the case of trimer, while two and three consecutive steps were possible in tetramer and pentamer hydrolyses, respectively. Chain lengthening increased catalytic efficiency (2.1 -> 8.6 M-1 s(-1)) and calculated kinetic constants showed a similarly increasing tendency (1.0 -> 6.7 x 10(-3) min(-1)). (C) 2012 Elsevier Ltd. All rights reserved.