Crystal Structures of Human Bifunctional Enzyme Aminoimidazole-4-carboxamide Ribonucleotide Transformylase/IMP Cyclohydrolase in Complex with Potent Sulfonyl-containing Antifolates

Cheom-Gil Cheong,Dennis W Wolan,Samantha E Greasley,Patricia A Horton,G Peter Beardsley,Ian A Wilson

doi:10.1074/jbc.m313691200

Cheom-Gil Cheong, Dennis W Wolan + Show 4 more

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https://doi.org/10.1074/jbc.m313691200

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Abstract

Aminoimidazole-4-carboxamide ribonucleotide (AICAR) transformylase/IMP cyclohydrolase (ATIC) is a bifunctional enzyme with folate-dependent AICAR transformylase and IMP cyclohydrolase activities that catalyzes the last two steps of purine biosynthesis. The AICAR transformylase inhibitors BW1540 and BW2315 are sulfamido-bridged 5,8-dideazafolate analogs with remarkably potent K(i) values of 8 and 6 nm, respectively, compared with most other antifolates. Crystal structures of ATIC at 2.55 and 2.60 A with each inhibitor, in the presence of substrate AICAR, revealed that the sulfonyl groups dominate inhibitor binding and orientation through interaction with the proposed oxyanion hole. These agents then appear to mimic the anionic transition state and now implicate Asn(431') in the reaction mechanism along with previously identified key catalytic residues Lys(266) and His(267). Potent and selective inhibition of the AICAR transformylase active site, compared with other folate-dependent enzymes, should therefore be pursued by further design of sulfonyl-containing antifolates.

Highlights

Aminoimidazole-4-carboxamide ribonucleotide (AICAR) transformylase/IMP cyclohydrolase (ATIC) is a bifunctional enzyme with folate-dependent AICAR transformylase and IMP cyclohydrolase activities that catalyzes the last two steps of purine biosynthesis
The crystal structures of human ATIC in complex with antifolates BW1540U88UD and BW2315U89UC provide the first view of how the substrate AICAR and folate-based co-factor moieties simultaneously occupy the AICAR Tfase active site
The refined human ATIC BW1540 structure consists of residues 4 –592 in monomer A and residues 5–592 in monomer B with 399 water molecules, two potassium ions, two BW1540 inhibitors within the AICAR Tfase domains, but only one xanthosine 5Ј-monophosphate (XMP) in the IMP cyclohydrolase active site of monomer A (Fig. 1B)

Summary

Introduction

Aminoimidazole-4-carboxamide ribonucleotide (AICAR) transformylase/IMP cyclohydrolase (ATIC) is a bifunctional enzyme with folate-dependent AICAR transformylase and IMP cyclohydrolase activities that catalyzes the last two steps of purine biosynthesis. Crystal structures of ATIC at 2.55 and 2.60 Å with each inhibitor, in the presence of substrate AICAR, revealed that the sulfonyl groups dominate inhibitor binding and orientation through interaction with the proposed oxyanion hole. These agents appear to mimic the anionic transition state and implicate Asn431؅ in the reaction mechanism along with previously identified key catalytic residues Lys266 and His267. Potent and selective inhibition of the AICAR transformylase active site, compared with other folate-dependent enzymes, should be pursued by further design of sulfonylcontaining antifolates. No evidence of channeling or tunneling of 5-formyl-AICAR between the two domains has been forthcoming [5, 8], strong electropositive patches that connect the two active sites may sequester 5-formylAICAR to the vicinity of the binding sites

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