Role of non-glycine residues in left-handed helical conformation for the conformational stability of human lysozyme

To understand the role of non-Gly residues in the left-handed helical confo rmation for the conformational stability of a protein, the non-Gly to Gly a nd Ala mutations at six left-handed residues (R21, Y38, R50, Q58, H78, and N118) of the human lysozyme were examined. The thermodynamic parameters for denaturation were determined using a differential scanning calorimeter, an d the crystal structures were analyzed by X-ray crystallography. If a left- handed non-Gly had an unfavorable steric interaction between the sidechain C beta and backbone, the Gly mutation would be expected to stabilize more t han the Ala mutation at the same position. For the mutant human lysozymes, however, there were few differences in the denaturation Gibbs energy (Delta G) between the Gly and Ala mutants, except for the substitution at position 58. Analysis of the changes in stability (Delta DeltaG) based on the struc tures of the wild-type and mutant proteins showed that the experimental Del ta DeltaG value of Q58G was similar to7 kJ/mol higher than the estimated va lue without consideration of any local steric interaction. These results in dicate that only Q58G increased the stability by elimination of local const raints. The residue 58 is located at the most rigid position in the left-ha nded non-Gly residues and is involved in its enzymatic function. It can be concluded that the left-handed non-Gly residues do not always have unfavora ble strain energies as compared with Gly at the same position. (C) 2001 Wil ey-Liss, Inc.