title: Binding of beta-D-glucopyranosyl bismethoxyphosphoramidate to glycogen phosphorylase b: kinetic and crystallographic studies.
abstract: In an attempt to identify a new lead molecule that would enable the design of inhibitors with enhanced affinity for glycogen phosphorylase (GP), beta-D-glucopyranosyl bismethoxyphosphoramidate (phosphoramidate), a glucosyl phosphate analogue, was tested for inhibition of the enzyme. Kinetic experiments showed that the compound was a weak competitive inhibitor of rabbit muscle GPb (with respect to alpha-D-glucose-1-phosphate (Glc-1-P)) with a Ki value of 5.9 (+/-0.1) mM. In order to elucidate the structural basis of inhibition, we determined the structure of GPb complexed with the phosphoramidate at 1.83 A resolution. The complex structure reveals that the inhibitor binds at the catalytic site and induces significant conformational changes in the vicinity of this site. In particular, the 280s loop (residues 282-287) shifts 0.4-4.3 A (main-chain atoms) to accommodate the phosphoramidate, but these conformational changes do not lead to increased contacts between the inhibitor and the protein that would improve ligand binding.
authors.affiliation: Institute of Organic and Pharmaceutical Chemistry, The National Hellenic Research Foundation, 48 Vassileos Constantinou Ave., 11635 Athens, Greece.
authors.forename: Evangelia D
authors.initials: ED
authors.lastname: Chrysina
authors.forename: Magda N
authors.initials: MN
authors.lastname: Kosmopoulou
authors.forename: Rozina
authors.initials: R
authors.lastname: Kardakaris
authors.forename: Nicolas
authors.initials: N
authors.lastname: Bischler
authors.forename: Demetres D
authors.initials: DD
authors.lastname: Leonidas
authors.forename: Thanukrishnan
authors.initials: T
authors.lastname: Kannan
authors.forename: Duraikkannu
authors.initials: D
authors.lastname: Loganathan
authors.forename: Nikos G
authors.initials: NG
authors.lastname: Oikonomakos
mesh_terms: D000577:Amides / Q000378:metabolism*; D020134:Catalytic Domain; D005947:Glucose / Q000031:analogs & derivatives* / Q000378:metabolism*; D024981:Glycogen Phosphorylase / Q000378:metabolism*; D007700:Kinetics; D008958:Models, Molecular; D015394:Molecular Structure
chemical_list: D000577:Amides; C502759:beta-D-glucopyranosyl bismethoxyphosphoramidate; D024981:Glycogen Phosphorylase; D005947:Glucose