N5-CAIR Mutase:  Role of a CO2 Binding Site and Substrate Movement in Catalysis,

Hoskins, Aaron A.; Morar, Mariya; Kappock, T.J.; Mathews, I.I.; Zaugg, Judith B.; Barder, Timothy E.; Peng, Paul; Okamoto, Akimitsu; Ealick, S.E.; Stubbe, JoAnne

doi:10.1021/bi602436g

Cited by 23 publications

(75 citation statements)

References 39 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In this way, the developing p orbital is geometrically disposed to achieve maximal overlap with the thiazolium π orbitals, thus lowering the barrier to bond breaking. This mechanism has been proposed for ThDP-dependent catalysis of decarboxylation (43)(44)(45). The present structure offers additional experimental corroboration of this model.…”

Section: Interaction With Iminium N4' Of Thdpsupporting

confidence: 86%

Probing the Active Center of Benzaldehyde Lyase with Substitutions and the Pseudosubstrate Analogue Benzoylphosphonic Acid Methyl Ester

et al. 2008

View full text Add to dashboard Cite

Benzaldehyde lyase (BAL) catalyzes the reversible cleavage of (R)-benzoin to benzaldehyde utilizing thiamin diphosphate and Mg 2+ as cofactors. The enzyme is important for the chemoenzymatic synthesis of a wide range of compounds via its carboligation reaction mechanism. In addition to its principal functions, BAL can slowly decarboxylate aromatic amino acids such as benzoylformic acid. It is also intriguing mechanistically due to the paucity of acid-base residues at the active center that can participate in proton transfer steps thought to be necessary for these types of reactions. Here methyl benzoylphosphonate, an excellent electrostatic analogue of benzoylformic acid, is used to probe the mechanism of benzaldehyde lyase. The structure of benzaldehyde lyase in its covalent complex with methyl benzoylphosphonate was determined to 2.49 Å (Protein Data Bank entry 3D7K) and represents the first structure of this enzyme with a compound bound in the active site. No large structural reorganization was detected compared to the complex of the enzyme with thiamin diphosphate. The configuration of the predecarboxylation thiamin-bound intermediate was clarified by the structure. Both spectroscopic and X-ray structural studies are consistent with inhibition resulting from the binding of MBP to the thiamin diphosphate in the active centers. We also delineated the role of His29 (the sole potential acid-base catalyst in the active site other than the highly conserved Glu50) and Trp163 in cofactor activation and catalysis by benzaldehyde lyase.

show abstract

Section: Interaction With Iminium N4' Of Thdpsupporting

confidence: 86%

Probing the Active Center of Benzaldehyde Lyase with Substitutions and the Pseudosubstrate Analogue Benzoylphosphonic Acid Methyl Ester

et al. 2008

View full text Add to dashboard Cite

show abstract

“…The second mechanism (path b) involves direct attack of CO 2 with assistance of the electron-rich exocyclic amine. The resulting tetrahedral intermediate, previously termed iso-CAIR, is then converted into CAIR via deprotonation of the C4 carbon6. As shown in figure 2B, NAIR could (in principle) have characteristics of a transition state for either mechanism.…”

Section: Introductionmentioning

confidence: 99%

Interrogating the mechanism of a tight binding inhibitor of AIR carboxylase

Firestine

Youn

et al. 2009

Bioorganic & Medicinal Chemistry

View full text Add to dashboard Cite

The enzyme aminoimidazole ribonucleotide (AIR) carboxylase catalyzes the synthesis of the purine intermediate, 4-carboxy-5-aminoimidazole ribonucleotide (CAIR). Previously, we have shown that the compound 4-nitro-5-aminoimidazole ribonucleotide (NAIR) is a slow, tight binding inhibitor of the enzyme with a K i of 0.34 nM. The structural attributes and the slow, tight binding characteristics of NAIR implicated this compound as a transition state or reactive intermediate analog. However, it is unclear what molecular features of NAIR contribute to the mimetic properties for either of the two proposed mechanisms of AIR carboxylase. In order to gain additional information regarding the mechanism for the potent inhibition of AIR carboxylase by NAIR, a series of heterocyclic analogs were prepared and evaluated. We find that all compounds are weaker inhibitors than NAIR and that CAIR analogs are not alternative substrates for the enzyme. Surprisingly, rather subtle changes in the structure of NAIR can lead to profound changes in binding affinity. Computational investigations of enzyme intermediates and these inhibitors reveal that NAIR displays an electrostatic potential surface similar to a proposed reaction intermediate. The result indicates that AIR carboxylase is likely sensitive to the electrostatic surface of reaction intermediates and thus compounds which mimic these surfaces should possess tight binding characteristics. Given the evolutionary relationship between AIR carboxylase and N 5 -CAIR mutase, we believe that this concept extends to the mutase enzyme as well. The implications of this hypothesis for the design of selective inhibitors of the N 5 -CAIR mutase are discussed.

show abstract

“…A conserved histidine residue is believed to serve as the acid and base for this reaction. 13 A channel runs through the center of the octamer along the fourfold-symmetry axis and we propose that this could be a relatively promiscuous binding site for small molecules (see Discussion below). The octameric structure, with its eight active sites, provided a distinct advantage in our molecular dynamics (MD) based fragment studies.…”

Section: Introductionmentioning

confidence: 93%

Fragment-Based Drug Discovery Using a Multidomain, Parallel MD-MM/PBSA Screening Protocol

Zhu

Lee

Lei

et al. 2013

J. Chem. Inf. Model.

View full text Add to dashboard Cite

We have developed a rigorous computational screening protocol to identify novel fragment-like inhibitors of N5-CAIR mutase (PurE), a key enzyme involved in de novo purine synthesis that represents a novel target for the design of antibacterial agents. This computational screening protocol utilizes molecular docking, graphics processing unit (GPU)-accelerated molecular dynamics and Molecular Mechanics/Poisson-Boltzmann Surface Area (MM/PBSA) free energy estimations to investigate the binding modes and energies of fragments in the active sites of PurE. PurE is a functional octamer comprised of identical subunits. The octameric structure, with its eight active sites, provided a distinct advantage in these studies because, for a given simulation length, we were able to place eight separate fragment compounds in the active sites to increase the throughput of the MM/PBSA analysis. To validate this protocol, we have screened an in-house fragment library consisting of 352 compounds. The theoretical results were then compared with the results of two experimental fragment screens, Nuclear Magnetic Resonance (NMR) and Surface Plasmon Resonance (SPR) binding analyses. In these validation studies, the protocol was able to effectively identify the competitive binders that had been independently identified by experimental testing, suggesting the potential utility of this method for the identification of novel fragments for future development as PurE inhibitors.

show abstract

N⁵-CAIR Mutase: Role of a CO₂ Binding Site and Substrate Movement in Catalysis^,

Cited by 23 publications

References 39 publications

Probing the Active Center of Benzaldehyde Lyase with Substitutions and the Pseudosubstrate Analogue Benzoylphosphonic Acid Methyl Ester

Probing the Active Center of Benzaldehyde Lyase with Substitutions and the Pseudosubstrate Analogue Benzoylphosphonic Acid Methyl Ester

Interrogating the mechanism of a tight binding inhibitor of AIR carboxylase

Fragment-Based Drug Discovery Using a Multidomain, Parallel MD-MM/PBSA Screening Protocol

Contact Info

Product

Resources

About