In order to improve the thermostability of α-L-rhamnosidase, four single site mutants: K406R, K440R, K573V, and E631F, were combined and four new mutants: K406R-K573V, K406R-E631F, K440R-K573V, and K440R-E631F, were obtained. In comparison to the wild type (WT), the thermostabilities of K440R-K573V and K440R-E631F improved, as their half-lives at 60 ℃ increased by 1.13-fold and 1.44-fold, respectively. Moreover, their half-lives were both 1.64 times higher than that of WT at 65 ℃, and 1.88-fold (K440R-K573V) and 1.68-fold (K440R-E631F) higher at 70 ℃. Based on circular dichroism, molecular dynamics and micro-structural analysis, it was found that mutant K440R-E631F in comparison to WT had increased internal hydrophobicity. Additionally, its contents of α-helix, 123456-turn and random coil also increased, which may be related to the increased thermostability of α-L-rhamnosidase.
LIU Xiaoqin
,
YANG Yan
,
WU Zheyu
,
GONG Jianye
,
LIU Jianan
,
LIAO Hui
,
LI Wenjing
,
LI Lijun
. Enhanced thermostability of α-L-rhamnosidase by multiple-site mutation[J]. Food and Fermentation Industries, 2019
, 45(6)
: 23
-29
.
DOI: 10.13995/j.cnki.11-1802/ts.018703
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