ACQUISITION OF NATIVE BETA-STRAND TOPOLOGY DURING THE RAPID COLLAPSE PHASE OF PROTEIN-FOLDING

The 98 residue C-terminal domain of the cell-surface receptor protein CD2 (CD2.D1) has a beta-sandwich fold belonging to the immunoglobulin superfamily but lacking the usual disulfide bridges. Kinetic studies o n the folding/unfolding of CD2.D1 reveal that folding proceeds through a rapidly formed intermediate state [Parker, M. J., & Clarke, A. R. ( 1997) Biochemistry 36, 5786-5794]. To characterize the structural prop erties of this intermediate we have performed a series of amide hydrog en exchange studies using the pH competition method, in which folding and exchange are initiated simultaneously. The complex beta-sheet topo logy of this molecule makes it an ideal object for examining the acqui sition of backbone hydrogen bonds made between sequence-local and sequ ence-distant segments of the chain during folding. The pattern of prot ected amides in the intermediate reveal that the essential features of the beta-sheet topology of CD2.D1 are defined early in the folding pa thway, before the development of intimate side chain interactions char acteristic of the native state. The results are discussed in light of current issues concerning the mechanistic relevance of kinetic protein folding intermediates.