MOLECULAR RECOGNITION AND PROTON-TRANSFER PROCESSES IN MALTODEXTRINPHOSPHORYLASE - AN FTIR STUDY

Maltodextrinphosphorylase (MDP) catalyses the degradation and in vitro elongation of polysaccharides. This enzyme is a pyridoxal-5'-phosphat e-dependent (PLP-dependent) protein. The phosphate group of PLP plays a decisive role in the catalytic mechanism. Two hydrogen-bonded system s are present at the active site of MDP. Using Fourier transform infra red (FTIR) difference spectroscopy, the pH dependence of these hydroge n-bonded systems are studied. On the basis of these results, the proto nation changes in the active centre of MDP when the substrate phosphat e glucose-1-methylenephosphonate is added are discussed. Two hydrogen- bonded chains are formed: (1) a chain connecting Lys 533-PLP and gluco se-1-methylenephosphonate; this chain shows large proton polarizabilit y due to collective proton motion, but is polarized in such a way that the substrate phosphate remains protonated; (2) a chain connecting Ly s 539, Tyr 538 and Glu 637; via this chain the excess proton of Lys 53 9 is shifted to Glu 637 and remains there; thus, in the binary complex , the excess proton of Lys 539 cannot be transferred to the phosphate group of the substrate. Finally, we discuss how, in the ternary comple x, the C-1-O-1 bond between the glycosyl residue and the phosphate gro up can be destabilized and split, and how the glycosyloxocarbonium ion so formed can be added to the polysaccharide chain. (C) 1997 Elsevier Science B.V.