The engineering of a more thermally stable lactate dehydrogenase by reduction of the area of a water-accessible hydrophobic surface.
A site-directed mutant of Bacillus stearothermophilus lactate dehydrogenase (lactate:NAD+ oxidoreductase, EC 1.1.1.27) has been engineered in which the conserved hydrophobic residue isoleucine-250 has been replaced by the more hydrophilic residue asparagine. This isoleucine forms a large part of a water-accessible, hydrophobic surface in the active site of the apo-enzyme which is covered by the B-face of the nicotinamide ring when coenzymes are bound. Reduction in the area of this hydrophobic surface results in the mutant tetramer being more thermally stable than the wild-type enzyme.
Wigley
D B
DB
Department of Biochemistry, University of Bristol, U.K.
Clarke
A R
AR
Dunn
C R
CR
Barstow
D A
DA
Atkinson
T
T
Chia
W N
WN
Muirhead
H
H
Holbrook
J J
JJ
D001665:Binding Sites; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D001411:Geobacillus stearothermophilus / Q000201:enzymology / Q000235:genetics; D007532:Isoleucine; D007770:L-Lactate Dehydrogenase; D009154:Mutation; D013696:Temperature
D007532:Isoleucine; D007770:L-Lactate Dehydrogenase