One of the strategies used to overcome the cooperativity of protein folding is to study fragments and peptides.We suggest to study the folding of a family of fragments of barstar, an inhibitor of the ribonuclease barnase, corresponding to its progressive elongaiion from the N-terminus. The objective is to study the development of slructure in a small protein, by diseccting it into N-terminal fragments of increasing length. These fragments will be created by cleavage of full length mutant protein for larger peptides and by chemical synthesis for small peptides. The introduction of Met residues by protein engineering will be used as cleavage sites for cyanogen bromide.We will analyze the ability of complementary fragments to associate and form a native-like structure, an association/folding study that could show the docking of preforrned segments, that constituted folding initiation sites.
The project would include the construction of small fragments that could mimic the interaction between barnase and barstar.