Rational Design, Synthesis, Evaluation, and Crystal Structure of a Potent Inhibitor of Human GAR Tfase: 10-(Trifluoroacetyl)-5,10-dideazaacyclic-5,6,7,8-tetrahydrofolic Acid,

Citations Over TimeTop 22% of 2003 papers

Abstract

Glycinamide ribonucleotide transformylase (GAR Tfase) has been the target of anti-neoplastic intervention for almost two decades. Here, we use a structure-based approach to design a novel folate analogue, 10-(trifluoroacetyl)-5,10-dideazaacyclic-5,6,7,8-tetrahydrofolic acid (10-CF(3)CO-DDACTHF, 1), which specifically inhibits recombinant human GAR Tfase (K(i) = 15 nM), but is inactive (K(i) > 100 microM) against other folate-dependent enzymes that have been examined. Moreover, compound 1 is a potent inhibitor of tumor cell proliferation (IC(50) = 16 nM, CCRF-CEM), which represents a 10-fold improvement over Lometrexol, a GAR Tfase inhibitor that has been in clinical trials. Thus, this folate analogue 1 is among the most potent and selective inhibitors known toward GAR Tfase. Contributing to its efficacious activity, compound 1 is effectively transported into the cell by the reduced folate carrier and intracellularly sequestered by polyglutamation. The crystal structure of human GAR Tfase with folate analogue 1 at 1.98 A resolution represents the first structure of any GAR Tfase to be determined with a cofactor or cofactor analogue without the presence of substrate. The folate-binding loop of residues 141-146, which is highly flexible in both Escherichia coli and unliganded human GAR Tfase structures, becomes highly ordered upon binding 1 in the folate-binding site. Computational docking of the natural cofactor into this and other apo or complexed structures provides a rational basis for modeling how the natural cofactor 10-formyltetrahydrofolic acid interacts with GAR Tfase, and suggests that this folate analogue-bound conformation represents the best template to date for inhibitor design.

Related Papers

• → Multi‐site substrate binding and interplay in barley α‐amylase 1(2008)24 cited
• → Methodology and Problems of Protein‐Ligand Docking: Case Study of Dihydroorotate Dehydrogenase, Thymidine Kinase, and Phosphodiesterase 4(2002)29 cited
• [Active site of trypsin and chymotrypsin; catalytic site and substrate binding site].(1970)
• → Organic Cofactors as Coenzymes(2006)
• → Crystallographic fragment binding studies of the Mycobacterium tuberculosis trifunctional enzyme suggest binding pockets for the tails of the acyl-CoA substrates at its active sites and a potential substrate channeling path between them(2024)