NDLI: Attempting to remove the substrate inhibition ofL-lactate dehydrogenase fromBacillus stearothermophilus by site-directed mutagenesis

Content Provider	Springer Nature Link
Author	Binay, Bariş Karagüler, Nevin Gül
Copyright Year	2007
Abstract	L-lactate dehydrogenase (LDH) catalyzes the interconversion of an oxoacid (pyruvate) and hydroxy-acid (lactate) using the NADH/NAD$^{+}$ pair as a redox cofactor. The enzyme has a commercial significance, as it can be used to produce chiral building blocks for the synthesis of key pharmaceuticals and agrochemicals. However, the substrate inhibition which is due to an abortive NAD$^{+}$-pyruvate complex reducing the steady state concentration of functional LDH limits its use in industry. This substrate inhibition can be overcome by weaking the binding of NAD$^{+}$.The conserved aspartic acid residue at position 38 was replaced by the longer basic arginine side chain (D38R) using PCR based overlap extension mutagenesis technique in the hope of weakening NAD$^{+}$-binding. The mutant gene was overexpressed in theEscherichia coli high-expression vector pKK223-3 in JM105 cells; then, the mutant protein was produced. Comparing the effect of substrate inhibition in the arginine-38 mutant with wild-type, substrate inhibition is decreased threefold.
Starting Page	265
Ending Page	272
Page Count	8
File Format	PDF
ISSN	02732289
Journal	Applied Biochemistry and Biotechnology
Volume Number	141
Issue Number	2-3
e-ISSN	15590291
Language	English
Publisher	Humana Press
Publisher Date	2007-01-01
Publisher Place	Totowa
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	Biotechnology Biochemistry
Content Type	Text
Resource Type	Article
Subject	Medicine Molecular Biology Environmental Engineering Biochemistry Bioengineering Applied Microbiology and Biotechnology Biotechnology

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