The role of the Val57 amino-acid residue in the hinge loop of the human cystatin C. Conformational studies of the beta2-L1-beta3 segments of wild-type human cystatin C and its mutants.

Human cystatin C (HCC) is one of the amyloidogenic proteins to be shown to oligomerize via a three-dimensional domain swapping mechanism. This process precedes the formation of a stable dimer and proceeds particularly easily in the case of the L68Q mutant. According to the proposed mechanism, dimerization of the HCC precedes conformational changes within the beta2 and beta3 strands. In this article, we present conformational studies, using circular dichroism and MD methods, of the beta2-L1-beta3 (His43-Thr72) fragment of the HCC involved in HCC dimer formation. We also carried out studies of the beta2-L1-beta3 peptide, in which the Val57 residue was replaced by residues promoting beta-turn structure formation (Asp, Asn, or Pro). The present study established that point mutation could modify the structure of the L1 loop in the beta-hairpin peptide. Our results showed that the L1 loop in the peptide excised from human cystatin C is broader than that in cystatin C. In the HCC protein, broadening of the L1 loop together with the unfavorable L68Q mutation in the hydrophobic pocket could be a force sufficient to cause the partial unfolding and then the opening of HCC or its L68Q mutant structure for further dimerization. We presume further that the Asp57 and Asn57 mutations in the L1 loop of HCC could stabilize the closed form of HCC, whereas the Pro57 mutation could lead to the opening of the HCC structure and then to dimer/oligomer formation. Copyright (c) 2009 Wiley Periodicals, Inc.