NDLI: Engineering a Disulfide Bond in the Lid Hinge Region of Rhizopus chinensis Lipase: Increased Thermostability and Altered Acyl Chain Length Specificity

Content Provider	PubMed Central
Author	Yu, Xiao-wei Tan, Nian-jiang Xiao, Rong Xu, Yan
Editor	Permyakov, Eugene A.
Copyright Year	2012
Abstract	The key to enzyme function is the maintenance of an appropriate balance between molecular stability and structural flexibility. The lid domain which is very important for “interfacial activation” is the most flexible part in the lipase structure. In this work, rational design was applied to explore the relationship between lid rigidity and lipase activity by introducing a disulfide bond in the hinge region of the lid, in the hope of improving the thermostability of R. chinensis lipase through stabilization of the lid domain without interfering with its catalytic performance. A disulfide bridge between F95C and F214C was introduced into the lipase from R. chinensis in the hinge region of the lid according to the prediction of the “Disulfide by Design” algorithm. The disulfide variant showed substantially improved thermostability with an eleven-fold increase in the t 1/2 value at 60°C and a 7°C increase of T m compared with the parent enzyme, probably contributed by the stabilization of the geometric structure of the lid region. The additional disulfide bond did not interfere with the catalytic rate (k cat) and the catalytic efficiency towards the short-chain fatty acid substrate, however, the catalytic efficiency of the disulfide variant towards pNPP decreased by 1.5-fold probably due to the block of the hydrophobic substrate channel by the disulfide bond. Furthermore, in the synthesis of fatty acid methyl esters, the maximum conversion rate by RCLCYS reached 95% which was 9% higher than that by RCL. This is the first report on improving the thermostability of the lipase from R. chinensis by introduction of a disulfide bond in the lid hinge region without compromising the catalytic rate.
Related Links	http://dx.doi.org/10.1371/journal.pone.0046388
Starting Page	46388
File Format	PDF
ISSN	19326203
e-ISSN	19326203
Journal	PLoS ONE
Issue Number	10
Volume Number	7
Language	English
Publisher	Public Library of Science
Publisher Date	2012-10-01
Access Restriction	Open
Rights Holder	Public Library of Science
Subject Keyword	Biochemistry, Genetics and Molecular Biology(all) Agricultural and Biological Sciences(all) Medicine(all) Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Multidisciplinary

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