STRUCTURE-FUNCTION ANALYSES OF THE SSC1P, MDJ1P, AND MGE1P SACCHAROMYCES-CEREVISIAE MITOCHONDRIAL PROTEINS IN ESCHERICHIA-COLI

The DnaK, DnaJ, and GrpE proteins of Escherichia coli have been univer sally conserved across the biological kingdoms and work together to co nstitute a highly efficient molecular chaperone machine. We have exami ned the extent of functional conservation of Saccharomyces cerevisiae Ssc1p, Mdj1p, and Mge1p by analyzing their ability to substitute for t heir corresponding E. coli homologs in vivo. We found that the express ion of yeast Mge1p, the GrpE homolog, allowed for the deletion of the otherwise essential grpE gene off. coli, albeit only up to 40 degrees C. The inability of Mge1p tea substitute for GrpE at very high tempera tures is consistent with our previous finding that it specifically fai led to stimulate DnaK's ATPase at such extreme conditions. In contrast to Mge1p, overexpression of Mdj1p, the DnaJ homolog, was lethal in E. coli. This toxicity was specifically relieved by mutations which affe cted the putative zinc binding region of Mdj1p. Overexpression of a tr uncated version of Mdj1p, containing the J- and Gly/Phe-rich domains, partially substituted for DnaJ function at high temperature. A chimeri c protein, consisting of the J domain of Mdj1p coupled to the rest of DnaJ, acted as a super-DnaJ protein, functioning even more efficiently than wild-type DnaJ. In contrast to the results with Mge1p and Mdj1p, both the expression and function of Ssc1p, the DnaK homolog, were sev erely compromised in E. coli. We were unable to demonstrate and functi onal complementation by Ssc1p, even when coexpressed with its Mdj1p co chaperone in E. coli.