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

DispersinB (DspB), a member of β-1,6-N-acetylglucosaminidase group of GH 20 glycoside hydrolases, catalyses the biofilm degradation of several human pathogenic microorganisms. DspB is a (β/α)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 β(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 nonreducing-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 × 10−3 min−1).

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► DispersinB catalysed hydrolysis reaction of β(1,6)-N-acetylglucosamine oligomers (DP 2–5) was studied.
► Catalytic constants for studied DispersinB-oligomer substrate reactions were calculated.
► Strictly nonreducing-end specificity was observed.
► Parallel reactions as a first step of oligomer substrate hydrolysis were first reported.