STRUCTURE OF A GLUTATHIONYLATED HUMAN LYSOZYME - A FOLDING INTERMEDIATE MIMIC IN THE FORMATION OF A DISULFIDE BOND

The three-dimensional structure of a mutant human lysozyme, C77A-a, in which the residue Cys77 is replaced by alanine, has been refined to a n R value of 0.125 using 8230 reflections in the resolution range 10.0 -1.8 Angstrom. It has been shown that C77A-a, in which the counterpart of Cys77 (Cys95) is modified with glutathione, has been shown to mimi c an intermediate in the formation of the disulfide bond Cys77-Cys95 d uring the folding of human lysozyme [Hayano, Inaka, Otsu, Taniyama, Mi ki, Matsushima & Kikuchi (1993). FEES Lett. 328, 203-208]. An earlier structure demonstrates that its overall structure is essentially ident ical to that of the wild-type protein and served as the starting model . The refined model includes atoms for all protein residues (1-130), 2 0 glutathione atoms and 113 water atoms. Further refinement shows more clearly the details of the protein, the bound glutathione molecule an d solvent structure. However, the main-chain folding and the atomic th ermal factors of the loop region from Thr70 to Leu79 were highly affec ted by the binding of the glutathione molecule, as compared with those of the wild-type protein. The bound glutathione shifted the main-chai n atoms from Val74 to Ala77 by more than 6.0 Angstrom, and the tempera ture factors of the atoms in the loop region were quite high (more tha n 40 Angstrom(2)), indicating that the backbone conformation of this r egion is highly flexible and that the loop region is not folded in the specific conformation observed in the wild-type protein. These result s strongly suggest that the loop structure in human lysozyme is folded later than the other regions of