Bisubstrate analog inhibitors of 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase: new lead exhibits a distinct binding mode.
6-Hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK), a key enzyme in the folate biosynthesis pathway catalyzing the pyrophosphoryl transfer from ATP to 6-hydroxymethyl-7,8-dihydropterin, is an attractive target for developing novel antimicrobial agents. Previously, we studied the mechanism of HPPK action, synthesized bisubstrate analog inhibitors by linking 6-hydroxymethylpterin to adenosine through phosphate groups, and developed a new generation of bisubstrate inhibitors by replacing the phosphate bridge with a piperidine-containing linkage. To further improve linker properties, we have synthesized a new compound, characterized its protein binding/inhibiting properties, and determined its structure in complex with HPPK. Surprisingly, this inhibitor exhibits a new binding mode in that the adenine base is flipped when compared to previously reported structures. Furthermore, the side chain of amino acid residue E77 is involved in protein-inhibitor interaction, forming hydrogen bonds with both 2' and 3' hydroxyl groups of the ribose moiety. Residue E77 is conserved among HPPK sequences, but interacts only indirectly with the bound MgATP via water molecules. Never observed before, the E77-ribose interaction is compatible only with the new inhibitor-binding mode. Therefore, this compound represents a new direction for further development.
Macromolecular Crystallography Laboratory, National Cancer Institute, Frederick, MD 21702, USA.
Genbin
G
Shi
Gary
G
Shaw
Yue
Y
Li
Yan
Y
Wu
Honggao
H
Yan
Xinhua
X
Ji
D000225:Adenine / Q000737:chemistry; D001665:Binding Sites; D018360:Crystallography, X-Ray; D017854:Diphosphotransferases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000138:chemical synthesis / Q000737:chemistry*; D006860:Hydrogen Bonding; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011622:Pterins / Q000138:chemical synthesis / Q000737:chemistry*; D013379:Substrate Specificity
D004791:Enzyme Inhibitors; D011622:Pterins; D017854:Diphosphotransferases; C019684:2-amino-4-hydroxy-6-hydroxymethyldihydropteridine pyrophosphokinase; D000225:Adenine