GLUTAMINE, ALANINE OR GLYCINE REPEATS INSERTED INTO THE LOOP OF A PROTEIN HAVE MINIMAL EFFECTS ON STABILITY AND FOLDING RATES

Natural proteins can contain flexible regions in their polypeptide cha in. We have investigated the effects of glycine, alanine and glutamine repeats on the stability and folding of a protein by inserting stretc hes of 7 to 13 residues into a suitable position in a model system, th e chymotrypsin inhibitor-2 (CI2). This folds by residues (1-40) dockin g with residues (41-64) to form a folding nucleus. The peptides GQ(4)G M, GQ(6)GM, GQ(8)GM, GQ(10)GM, GA(2)SA(4)SA(2)GM and G(3)SG(4)SG(3)M w ere inserted after residue 40. The stability of the mutant proteins ch anges only weakly with chain length and nature of insertion, suggestin g that the presence of unstructured polypeptide chains in a protein do es not have a great energetic penalty. This has implications in cataly sis, for example, where floppy regions have been noted in active sites , and in DNA transcription where activators, transcription factors and intermediary proteins all show long repeats of glycine/serine and/or glutamine, which are thought to be important for function. We find tha t the rate of folding is very insensitive to the length of the linker. The changes in rate are close to these predicted from polymer theory for the loss of configuration entropy on closing a loop. This implies that all the diffusion steps are relatively rapid. (C) 1997 Academic P ress Limited.