Lid hinge region of Penicillium expansum lipase affects enzyme activity and interfacial activation

Lid hinge region of Penicillium expansum lipase affects enzyme activity and interfacial activation

•Sites displaying the highest B factors in the lid were selected for mutagenesis.•Mutants showed properties remarkably different from those of wild-type PEL.•Mutation shifted the chain-length specificity.•One mutant lacked interfacial activation.•We provide a new clue for selecting critical amino ac...

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Bibliographic Details
Published in:Process biochemistry (1991) Vol. 50; no. 8; pp. 1218 - 1223
Main Authors: Tang, Lianghua, Su, Min, Yan, Junzhe, Xie, Sheng, Zhang, Wenhuang
Format: Journal Article
Language:English
Published: Elsevier Ltd 01-08-2015
Subjects:
Activation
Amino acids
Biochemistry
Hinges
Lid hinge
Lipase
Mutagenesis
Penicillium
Penicillium expansum
Residues
Saturation
Penicillium expansum
Lid hinge
Mutagenesis
Lipase
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Summary:•Sites displaying the highest B factors in the lid were selected for mutagenesis.•Mutants showed properties remarkably different from those of wild-type PEL.•Mutation shifted the chain-length specificity.•One mutant lacked interfacial activation.•We provide a new clue for selecting critical amino acid residues for the enzyme design. Saturation mutagenesis at sites displaying the highest B factors in the lid and the hinge regions of Penicillium expansum lipase (PEL) has been employed to improve the efficiency of the lipase in biocatalysis. Replacements of amino acid on beneficial mutants were identified as T66L/D70N, T66V/D70N, E83K, E83H and E83N. In substrate specificity assays, T66L/D70N was significantly more active than wild-type PEL on substrates with medium and long chain lengths. In addition this mutant also displayed a 136.4-fold increase in activity on p-nitrophenyl palmitate. Remarkably, E83K lacked interfacial activation while it was observed in wild-type PEL and the other mutants. Insight into the relation between the mutations and enzymatic properties was gained by modeling and docking studies. All these mutants showed an enhanced catalytic activity, indicating their potential in further application. Therefore, these results indicate the amino acid composition of the lid hinge region plays an extremely important role in the interfacial activation, activity and substrate specificity of PEL. Moreover, the results in this work provide a new clue for selecting critical amino acid residues for the enzyme design.
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ISSN:1359-5113
1873-3298
DOI:10.1016/j.procbio.2015.04.022