Cell-free synthesis of watermelon glyoxysomal malate dehydrogenase: a comparison with the mitochondrial isoenzyme.

Cotyledons of dark-grown watermelon seedlings contain during the first period of germination, 4 and later 5 MDH isoenzymes. Whereas isoenzymes I, II, and IV belong to the cytosol, isoenzyme III is confined to the mitochondria (mMDH) and isoenzyme V to the glyoxysomes (gMDH). The organelle-bound MDHs were purified to homogeneity and characterized. They are remarkably similar with respect to their kinetic properties, but differ widely in other characteristics, e.g. MW, IEP, thermostability, and serological properties. Both isoenzymes are synthesized de novo during germination by cytoplasmic ribosomes. In a reticulocyte cell-free system programmed by watermelon poly (A+) RNA, gMDH and mMDH are synthesized as larger precursors as compared to the authentic polypeptide chains, with an extra sequence of ca. 8.0 kilodaltons in the case of gMDH and ca. 3.3 kilodaltons with mMDH. The processing of the gMDH precursor by an enriched organelle fraction in not complete. If the post-translational incorporation of the in vitro product into the organelles is followed by proteinase treatment that completely hydrolyzes noncompartmentalized gMDH, the authentic product is obtained. A model for the transport mechanism is given.