Hydrolysis of cyclic poly(ethylene terephthalate) trimers by a carboxylesterase from Thermobifida fusca KW3
We have identified a carboxylesterase produced in liquid cultures of the thermophilic actinomycete Thermobifida fusca KW3 that were supplemented with poly(ethylene terephthalate) fibers. The enzyme hydrolyzed highly hydrophobic, synthetic cyclic poly(ethylene terephthalate) trimers with an optimal activity at 60°C and a pH of 6. V max and K m values for the hydrolysis were 9.3 µmol−1 min−1 mg−1 and 0.5 mM, respectively. The esterase showed high specificity towards short and middle chain-length fatty acyl esters of p-nitrophenol. The enzyme retained 37% of its activity after 96 h of incubation at 50°C and a pH of 8. Enzyme inhibition studies and analysis of substitution mutants of the carboxylesterase revealed the typical catalytic mechanism of a serine hydrolase with a catalytic triad composed of serine, glutamic acid, and histidine.
KeywordsThermobifida fusca Carboxylesterase Cutinase Poly(ethylene terephthalate) (PET) Enzymatic hydrolysis
S. Billig was supported by grant no. 20004/730, Deutsche Bundesstiftung Umwelt and T. Oeser by grant no. 13-8811.61/215-1, Sächsisches Staatsministerium für Umwelt und Landwirtschaft. C. Roth from the University of Leipzig is acknowledged for his assistance in the pI determination. I. Neundorf and J. Stichel from the University of Leipzig are acknowledged for the amino acid sequence determination and MALDI-TOF/TOF analysis.
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