Unusual conformational changes in 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase as revealed by X-ray crystallography and NMR

The crystal structure of Escherichia coli 6-hydroxy-methyl-7,8-dihydropteri n pyrophosphokinase (HPPK) in complex with MgADP has been determined at 1.5 -Angstrom resolution with a crystallographic R factor of 0.191. The solutio n structure of HPPK in complex with Mg2+ and beta,gamma -methyleneadenosine 5'-triphosphate (MgAMPPCP) has been determined using a simulated annealing pro. tocol with 3,523 experimental NMR restraints. The root mean square de viation of the ensemble of 20 refined conformers that represent the solutio n structure from the mean coordinate set derived from them is 0.74 +/- 0.26 A for all backbone atoms and 0.49 +/- 0.22 Angstrom when residues Pro(14), Pro(44)-Gln(50), and Arg(84)-Pro(91) are excluded. Binding of MgADP causes significant changes in the conformation and dynamical property of three lo ops of HPPK that are involved in catalysis. A dramatic, unusual conformatio nal change is that loop 3 moves away from the active center significantly w ith some residues moving by > 17 Angstrom. The binding of MgADP also stabil izes loop 1 and loop 3 but makes loop 2 more mobile. Very similar conformat ional and dynamical changes are observed in the NAM solution structure of H PPK(.)MgAMPPCP. The conformational and dynamical changes may play important roles in both substrate binding and product release in the catalytic cycle .