STRUCTURAL AND FUNCTIONAL-PROPERTIES OF THE 34-KDA FRAGMENT PRODUCED BY THE N-TERMINAL CHYMOTRYPTIC CLEAVAGE OF GLUTATHIONE TRANSFERASE P1-1

Limited proteolysis of glutathione transferase P1-1 (GSTP1-1) by chymo trypsin generates a 34-kDa GSTP1-1 fragment (a dimer of the 17-kDa sub unit composed by residues 48-207) containing the whole C-terminal doma in and a part (about 15%) of the N-terminal domain (residues 48-76, i. e., the structural elements beta 3, beta 4, and alpha C). The structur al and functional properties of this large fragment have been investig ated by analyzing its binding properties to 2-p-toluidinylnaphthalene- 6-sulfonate (TNS) extrinsic probe, the TNS displacement technique, and the molecular modeling approach. The results obtained indicated that the 34-kDa GSTP1-1 fragment maintains an hydrophobic pocket with the s ame structural properties of the corresponding GSTP1-1 hydrophobic bin ding site. In addition, the 34-kDa GSTP1-1 binds a number of hydrophob ic compounds such as 1-chloro-2,4-dinitrobenzene, hemin, and bilirubin with the same affinity of the native enzyme. Being structurally and f unctionally autonomous, this fragment, mostly constituted by domain II , appears as an independent folding unit in the protein. Nevertheless, in the entire native protein, interdomain interactions occur and are responsible for the major solvent exposure of the H-site in the presen ce of glutathione. (C) 1995 Academic Press, Inc.