The three-dimensional structure of the recombinant Mgelp protein, which is a first mitochondrial homologue of the E. Coli GrpE protein, will be determined by the multidimensional heteronuclear NMR spectroscopy using the 15 N and 13 C labelled protein samples.
Mgelp consists of 196 amino acids and belongs to the heat shock proteins of the Hsp70 chaperone family that is responsible for folding and assembly of proteins, transcription, protection against heat, and translocation of proteins across intracellular membranes. The dynamic properties of the protein will be obtained from the 15 N and 13 C relaxation measurements. The binding site of Mgelp with mt-Hsp70 and its smaller peptide fragments will be mapped by monitoring perturbation of line widths and chemical shifts of the 15 N labelled Mgelp resonances during titration with unlabeled mt-Hsp70 and its peptides. The change of the conformational flexibility of the binding regions upon complexation with small peptides will also be studied.
As no structural information exists on the Mgelp and GrpE proteins, the proposed study, together with biochemical experiments, should serve as foundation for a better understanding of these protein-protein interaction .