Characterization of chimeric isocitrate dehydrogenases of a mesophilic nitrogen-fixing bacterium, Azotobacter vinelandii, and a psychrophilic bacterium, Colwellia maris.
Several properties of chimeric enzymes between a mesophilic isocitrate dehydrogenase (IDH) from a nitrogen-fixing bacterium, Azotobacter vinelandii, and a cold-adapted IDH isozyme (IDH-II) from a psychrophilic bacterium, Colwellia maris, were examined. Each of the genes encoding the IDHs was divided into four regions of almost equal lengths, and each region was ligated with different combinations to construct various chimeric genes. The resultant wild-type and chimeric genes were overexpressed in Escherichia coli. The wild-type and chimeric IDHs were classified into three groups based on optimum temperatures for activity of 20 degrees, 30 degrees, and 40 degrees C. The IDHs with a lower optimum temperature were more thermolabile. The optimum temperature and thermostability of the chimeric enzymes decreased on increasing the proportion derived from the cold-adapted IDH-II of C. maris. Furthermore, the C-terminal region of the C. maris IDH-II was suggested to be responsible for its psychrophilic characteristics.
Yoneta
Mizuho
M
Division of Biological Sciences, Graduate School of Science, Hokkaido University, Sapporo, Hokkaido 060-0810, Japan.
Sahara
Takehiko
T
Nitta
Katsutoshi
K
Takada
Yasuhiro
Y
D034241:Alteromonadaceae / Q000201:enzymology / Q000235:genetics*; D016948:Azotobacter vinelandii / Q000201:enzymology / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D007521:Isocitrate Dehydrogenase / Q000235:genetics* / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D013696:Temperature; D014169:Transformation, Bacterial
D001426:Bacterial Proteins; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D007521:Isocitrate Dehydrogenase