XVI International Botanical Congess

The two most N-terminal and the two most C-terminal cysteines in chloroplast malate dehydrogenase are present as disulfides in the crystal structure of the enzyme. But homology modeling suggests another disulfide involving the penultimate Cys residue and one of the internal Cys residues. Here we show that, unlike the wildtype maize enzyme, mutants lacking the penultimate Cys residue or that internal Cys residue are activated by mercaptoethanol. Replacement of the more C-terminal Cys results in an enzyme completely activated by mercaptoethanol. Replacement of the internal Cys residue results in an enzyme only partially activated by mercaptoethanol. We suggest that the penultimate Cys residue undergoes continual thiol-disulfide exchange between the last Cys residue in the C-terminus and the internal Cys residue.