CYSTEINE SYNTHESIS IN PLANTS - PROTEIN-PROTEIN INTERACTIONS OF SERINEACETYLTRANSFERASE FROM ARABIDOPSIS-THALIANA

The biosynthesis of cysteine represents the final step of sulfate assi milation in bacteria and plants. It is catalyzed by the sequential act ion of serine acetyltransferase (SAT) and O-acetylserine (thiol) lyase (OAS-TL) which form a cysteine synthase (CS) complex in vitro. SAT an d OAS-TL from Arabidopsis thaliana have previously been cloned, and no w the first evidence is presented for the CS complex and SAT self-inte raction in vivo employing the yeast two-hybrid system. Application of this method proved to be an efficient tool for the analysis of protein -protein interactions within a plant metabolic protein complex. Mappin g of SAT domain structure revealed two new, independent domains with s pecific functions in protein-protein interaction. Analysis using trunc ated proteins proved the C-terminus of SAT to be sufficient for associ ation with OAS-TL and to correlate with the putative transferase activ ity domain. SAT/SAT interaction was localized in the central region of the protein and occured also between SAT isoforms. Both protein inter action domains coincided with distinct alpha-helical and beta-sheet cl usters and together correlated with the minimal protein structure requ ired for SAT catalysis as shown by functional complementation of an Es cherichia coli mutant. The homo- and hetero-oligomerization properties are discussed with respect to the assumed function of the CS complex in metabolic channeling and activation of SAT by interaction with OAS- TL.