CRYSTAL-STRUCTURES AND SOLUTION CONFORMATIONS OF A DOMINANT-NEGATIVE MUTANT OF ESCHERICHIA-COLI MALTOSE-BINDING PROTEIN

A mutant of the periplasmic maltose-binding protein (MBP) with altered transport properties was studied. A change of residue 230 from trypto phan to arginine results in dominant-negative MBP: expression of this protein against a wild-type background causes inhibition of maltose tr ansport. As part of an investigation of the mechanism of such inhibiti on, we have solved crystal structures of both unliganded and liganded mutant protein. In the closed, liganded conformation, the side-chain o f R230 projects into a region of the surface of MBP that has been iden tified as important for transport while in the open form, the same sid e-chain takes on a different, and less ordered, conformation. The crys tallographic work is supplemented with a small-angle X-ray scattering study that provides evidence that the solution conformation of unligan ded mutant is similar to that of wild-type MBP. It is concluded that d ominant-negative inhibition of maltose transport must result from the formation of a non-productive complex between liganded-bound mutant MB P and wild-type MalFGK(2). A general kinetic framework for transport b y either wild-type MalFGK(2) or MBP-independent MaIFGK(2) is used to u nderstand the effects of dominant-negative MBP molecules on both of th ese systems. (C) 1996 Academic Press Limited