Binding of beta-D-glucopyranosyl bismethoxyphosphoramidate to glycogen phosphorylase b: kinetic and crystallographic studies.
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.
Chrysina
Evangelia D
ED
Institute of Organic and Pharmaceutical Chemistry, The National Hellenic Research Foundation, 48 Vassileos Constantinou Ave., 11635 Athens, Greece.
Kosmopoulou
Magda N
MN
Kardakaris
Rozina
R
Bischler
Nicolas
N
Leonidas
Demetres D
DD
Kannan
Thanukrishnan
T
Loganathan
Duraikkannu
D
Oikonomakos
Nikos G
NG
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
D000577:Amides; C502759:beta-D-glucopyranosyl bismethoxyphosphoramidate; D024981:Glycogen Phosphorylase; D005947:Glucose