Ks. Park et Ml. Kahn, Distribution of two isoforms of NADP-dependent isocitrate dehydrogenase insoybean (Glycine max), PLANT MOL B, 40(1), 1999, pp. 13-21
Two different cDNAs that encode NADP-specific isocitrate dehydrogenase (NAD
P-IDH) isozymes of soybean (Glycine max) were characterized. The nucleotide
sequences of the coding regions of these cDNAs have 74% identity to each o
ther and give predicted amino acid sequences that have 83% identity to each
other. Using PCR techniques, their coding regions were subcloned into a pr
otein overexpression vector, pQE32, to yield pIDH4 and pIDH1, respectively.
Both IDH4 and IDH1 enzymes were expressed in Escherichia coli as catalytic
ally active His6 tagged proteins, purified to homogeneity by affinity chrom
atography on nickel chelate resin and rabbit polyclonal antibodies to each
were generated. Surprisingly, antiserum to IDH4 did not react with IDH1 pro
tein and IDH1 antiserum reacted only very weakly with IDH4 protein. IDH4 an
tibody reacts with a protein of expected molecular weight in cotyledon, you
ng leaf, young root, mature root and nodules but the reaction with mature l
eaf tissue was low compared to other tissues. Western blot results show tha
t IDH1 was not expressed in young roots but a protein that reacts with the
IDH1 antibody was highly expressed in leaves, showing that there was tissue
-specific accumulation of NADP-IDH isozymes in soybean.