Multi-targeted antifolates aimed at avoiding drug resistance form covalent closed inhibitory complexes with human and Escherichia coli thymidylate synthases

Crystal structures of four pyrrolo(2,3-d)pyrimidine-based antifolate compounds, developed as inhibitors of thymidylate synthase (TS) in a strategy to circumvent drug-resistance, have been determined in complexes with their in vivo target, human thymidylate synthase, and with the structurally best-ch...

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Bibliographic Details
Published in:Journal of molecular biology 2001-11, Vol.313 (4), p.813-829
Main Authors: Sayre, Peter H, Finer-Moore, Janet S, Fritz, Timothy A, Biermann, Donna, Gates, Susan B, MacKellar, Warren C, Patel, Vinod F, Stroud, Robert M
Format: Article
Language:English
Subjects:
antifolate
Apoenzymes - chemistry
Apoenzymes - genetics
Apoenzymes - metabolism
Binding Sites
cancer
Crystallization
Crystallography, X-Ray
Deoxyuracil Nucleotides - metabolism
Dimerization
drug resistance
Drug Resistance - genetics
dTMP synthesis
Enzyme Inhibitors - chemistry
Enzyme Inhibitors - metabolism
Enzyme Inhibitors - pharmacology
Escherichia coli
Escherichia coli - enzymology
Folic Acid Antagonists - chemistry
Folic Acid Antagonists - metabolism
Folic Acid Antagonists - pharmacology
Glutamates - chemistry
Glutamates - metabolism
Glutamates - pharmacology
Guanine - analogs & derivatives
Guanine - chemistry
Guanine - metabolism
Guanine - pharmacology
Humans
Hydrogen Bonding
Ligands
LY231514
Models, Molecular
Mutation - genetics
Pemetrexed
Peptide Synthases - metabolism
Protein Conformation - drug effects
Protein Folding
Static Electricity
Thymidylate Synthase - antagonists & inhibitors
Thymidylate Synthase - chemistry
Thymidylate Synthase - genetics
Thymidylate Synthase - metabolism
X-ray crystal structure
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Summary:Crystal structures of four pyrrolo(2,3-d)pyrimidine-based antifolate compounds, developed as inhibitors of thymidylate synthase (TS) in a strategy to circumvent drug-resistance, have been determined in complexes with their in vivo target, human thymidylate synthase, and with the structurally best-characterized Escherichia coli enzyme, to resolutions of 2.2-3.0 Å. The 2.9 Å crystal structure of a complex of human TS with one of the inhibitors, the multi-targeted antifolate LY231514, demonstrates that this compound induces a “closed” enzyme conformation and leads to formation of a covalent bond between enzyme and substrate. This structure is one of the first liganded human TS structures, and its solution was aided by mutation to facilitate crystallization. Structures of three other pyrrolo(2,3-d)pyrimidine-based antifolates in complex with Escherichia coli TS confirm the orientation of this class of inhibitors in the active site. Specific interactions between the polyglutamyl moiety and a positively charged groove on the enzyme surface explain the marked increase in affinity of the pyrrolo(2,3-d)pyrimidine inhibitors once they are polyglutamylated, as mediated in vivo by the cellular enzyme folyl polyglutamate synthetase.
ISSN:0022-2836
1089-8638
DOI:10.1006/jmbi.2001.5074