TY - JOUR
T1 - Improvement of the activity of arylmalonate decarboxylase by random mutagenesis
AU - Terao, Y.
AU - Miyamoto, K.
AU - Ohta, H.
N1 - Funding Information:
Acknowledgements This research was supported in part by the Ministry of Education, Culture, Sports, Science and Technology, Grant-in-Aid for the 21st Century Center of Excellence Program entitled “Understanding and Control of Life’s Function via Systems Biology (Keio University).” Financial support from Takeda Science Foundation is also greatly acknowledged.
PY - 2006/12
Y1 - 2006/12
N2 - Arylmalonate decarboxylase (EC 4.1.1.76) catalyzes enantioselective decarboxylation of α-aryl-α-methylmalonates to give optically pure α-arylpropionates. Recently, we have succeeded in creating a double mutant enzyme that gave opposite enantionmer as the product. Unfortunately, however, the activity of the mutant decreased far lower than that of the native enzyme. Thus, we performed the directed evolution of the mutant via the random mutagenesis method employing the mutator strain Escherichia coli XL1-Red. About 50,000 mutants were screened on color assay plate, and one mutant with higher activity was obtained. Gene analysis of this mutant indicated that the obtained enzyme had an S36N mutation in addition to its original G74C/C188S mutations. The activity of the triple mutant enzyme was tenfold higher than that of the starting doubly mutated enzyme.
AB - Arylmalonate decarboxylase (EC 4.1.1.76) catalyzes enantioselective decarboxylation of α-aryl-α-methylmalonates to give optically pure α-arylpropionates. Recently, we have succeeded in creating a double mutant enzyme that gave opposite enantionmer as the product. Unfortunately, however, the activity of the mutant decreased far lower than that of the native enzyme. Thus, we performed the directed evolution of the mutant via the random mutagenesis method employing the mutator strain Escherichia coli XL1-Red. About 50,000 mutants were screened on color assay plate, and one mutant with higher activity was obtained. Gene analysis of this mutant indicated that the obtained enzyme had an S36N mutation in addition to its original G74C/C188S mutations. The activity of the triple mutant enzyme was tenfold higher than that of the starting doubly mutated enzyme.
KW - Activity
KW - Arylmalonate decarboxylase
KW - Random-mutagenesis
KW - XL1-Red
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U2 - 10.1007/s00253-006-0518-z
DO - 10.1007/s00253-006-0518-z
M3 - Article
C2 - 16865343
AN - SCOPUS:33751021095
SN - 0175-7598
VL - 73
SP - 647
EP - 653
JO - Applied Microbiology and Biotechnology
JF - Applied Microbiology and Biotechnology
IS - 3
ER -