The importance of loop length in the folding of an immunoglobulin domain

Abstract

Immunoglobulin (Ig)-like proteins have been shown to fold following formation of a nucleus comprising interactions between residues that are distant in the primary sequence. What role do the loops connecting these nucleus residues play? Here, the importance of loops connecting β-strands in different sheets of the Ig fold is investigated, by insertion of five glycine residues into the B-C loop of an Ig domain from human titin, TI I27. The folding pathway of this elongated 'pseudo wild-type' TI I27 is probed using protein engineering and Φ-value analysis. The Φ-values calculated for mutants within the pseudo wild-type protein indicate that the folding nucleus in wild-type TI I27 is conserved, supporting the hypothesis that the inter-sheet loop is not critical to the formation of a long-range folding nucleus.