Developmental expression of three mungbean Hsc70s and substrate-binding specificity of the encoded proteins

We isolated three mungbean Hsc70 cDNAs (VrHsc70-1, 70-2 and 70-3) and characterized their developmental expression at both the transcript and protein levels. We also characterized the binding specificity between each VrHsc70 protein and its potential substrates. RNase protection assays showed that these three cytosolic VrHsc70 genes were expressed similarly in all organs at all times during the mungbean life cycle, except at the initiation of germination and during late seed embryogenesis. Western blotting analyses showed that a different group of cytosolic VrHsc70 proteins accumulated in dehydrated seeds during seed maturation and the accumulated proteins remained high during the early stages of germination. Binding specificities of these three mungbean Hsc70s were determined using the C-terminal 30 kDa of the three VrHsc70s to select bound heptapeptides using phage display screening, and were confirmed by ELISA. We found that the heptapeptides, KVWVLPI, KLWVIPQ and YAPLSRL, specifically bound to the C-terminal 30 kDa region of VrHsc70-1, 70-2 and 70-3, respectively. The hydrophobic residues in the core of the heptapeptides, as well as residues 6 and 7, might contribute to the binding specificity. Our results indicate that the function of these three VrHsc70s may not be important in seed maturation or in desiccation tolerance, but are more likely involved in normal growth and development.