<i>Escherichia coli</i> dihydrodipicolinate synthase: characterization of the imine intermediate and the product of bromopyruvate treatment by electrospray mass spectrometry

Borthwick, Emma B.; Connell, Susanne J.; Tudor, David W.; Robins, David J.; Shneier, Andrea; Abell, Chris; Coggins, John R.

doi:10.1042/bj3050521

Cited by 43 publications

(29 citation statements)

References 17 publications

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“…This has been confirmed by sodium borohydride trapping experiments, 4 and the X-ray crystal structure of DHDPS with pyruvate bound. 3 Subsequent tautomerisation gives the enamine 5.…”

supporting

confidence: 68%

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Two new irreversible inhibitors of dihydrodipicolinate synthase: diethyl (E,E)-4-oxo-2,5-heptadienedioate and diethyl (E)-4-oxo-2-heptenedioate

Turner

Healy

Dobson

et al. 2005

Bioorganic & Medicinal Chemistry Letters

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“…This has been confirmed by sodium borohydride trapping experiments, 4 and the X-ray crystal structure of DHDPS with pyruvate bound. 3 Subsequent tautomerisation gives the enamine 5.…”

supporting

confidence: 68%

“…DHDPS catalyses the condensation of (S)-aspartate semialdehyde (ASA, 2) 2 and pyruvate (1) to form an unstable heterocycle, formerly thought to be dihydrodipicolinate (4), but now believed to be 4-hydroxytetrahydrodipicolinate (3), 3 with spontaneous dehydration to give 4 following release from the enzyme active site (Fig. 1).…”

mentioning

confidence: 99%

Two new irreversible inhibitors of dihydrodipicolinate synthase: diethyl (E,E)-4-oxo-2,5-heptadienedioate and diethyl (E)-4-oxo-2-heptenedioate

Turner

Healy

Dobson

et al. 2005

Bioorganic & Medicinal Chemistry Letters

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“…Since the first crystal structure was reported in 1995 [5], Escherichia coli DHDPS (EcDHDPS) has been used extensively as a model for the reaction catalyzing the condensation of (S)-aspartate-␤-semialdehyde ((S)-ASA) and pyruvate to form an unstable heterocyclic product, currently thought to be (4S)-4-dydroxy-2,3,4,5-tetrahydro-(2S)-dipicolinic acid (HTPA) [6][7][8][9][10]. Complex structures and subsequent mutagenesis studies proposed that three residues Tyr133, Thr44 and Tyr107, act as a proton shuttle to transfer protons to and from the active site [8,[11][12][13][14].…”

Section: Introductionmentioning

confidence: 99%

Crystal structure of dihydrodipicolinate synthase from Hahella chejuensis at 1.5 Å resolution

Kang

Kim

Ahn

et al. 2010

International Journal of Biological Macromolecules

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“…However, from these data, we cannot preclude the binding of b-hydroxypyruvate to the enzyme at locations other than at the active site (see the following data from mass spectrometry and the X-ray model). Another pyruvate analog, 3-bromopyruvate, is a known irreversible inhibitor of DHDPS (Laber et al 1992), although it does not specifically alkylate residues at the active site (Borthwick et al 1995).…”

Section: B-hydroxypyruvate Is a Time-dependent Inhibitor Of Dhdpsmentioning

confidence: 99%

Conserved main‐chain peptide distortions: A proposed role for Ile203 in catalysis by dihydrodipicolinate synthase

et al. 2008

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In recent years, dihydrodipicolinate synthase (DHDPS, E.C. 4.2.1.52) has received considerable attention from a mechanistic and structural viewpoint. DHDPS catalyzes the reaction of (S)-aspartate-b-semialdehyde with pyruvate, which is bound via a Schiff base to a conserved active-site lysine (Lys161 in the enzyme from Escherichia coli). To probe the mechanism of DHDPS, we have studied the inhibition of E. coli DHDPS by the substrate analog, b-hydroxypyruvate. The K i was determined to be 0.21 (60.02) mM, similar to that of the allosteric inhibitor, (S)-lysine, and b-hydroxypyruvate was observed to cause timedependent inhibition. The inhibitory reaction with b-hydroxypyruvate could be qualitatively followed by mass spectrometry, which showed initial noncovalent adduct formation, followed by the slow formation of the covalent adduct. It is unclear whether b-hydroxypyruvate plays a role in regulating the biosynthesis of meso-diaminopimelate and (S)-lysine in E. coli, although we note that it is present in vivo. The crystal structure of DHDPS complexed with b-hydroxypyruvate was solved. The active site clearly showed the presence of the inhibitor covalently bound to the Lys161. Interestingly, the hydroxyl group of bhydroxypyruvate was hydrogen-bonded to the main-chain carbonyl of Ile203. This provides insight into the possible catalytic role played by this peptide unit, which has a highly strained torsion angle (v ;201°). A survey of the known DHDPS structures from other organisms shows this distortion to be a highly conserved feature of the DHDPS active site, and we propose that this peptide unit plays a critical role in catalysis.Keywords: dihydrodipicolinate synthase; lysine biosynthesis; conserved peptide distortion; X-ray crystallography Abbreviations: HTPA, (4S)-4-hydroxy-2,3,4,5-tetrahydro-(2S)-dipicolinic acid; DHDPS, dihydrodipicolinate synthase; DHDPR, dihydrodipicolinate reductase; (S)-ASA, (S)-aspartate-bsemialdehyde; DAP, meso-diaminopimelate; MS, mass spectrometry.Article and publication are at

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Escherichia coli dihydrodipicolinate synthase: characterization of the imine intermediate and the product of bromopyruvate treatment by electrospray mass spectrometry

Cited by 43 publications

References 17 publications

Two new irreversible inhibitors of dihydrodipicolinate synthase: diethyl (E,E)-4-oxo-2,5-heptadienedioate and diethyl (E)-4-oxo-2-heptenedioate

Two new irreversible inhibitors of dihydrodipicolinate synthase: diethyl (E,E)-4-oxo-2,5-heptadienedioate and diethyl (E)-4-oxo-2-heptenedioate

Crystal structure of dihydrodipicolinate synthase from Hahella chejuensis at 1.5 Å resolution

Conserved main‐chain peptide distortions: A proposed role for Ile203 in catalysis by dihydrodipicolinate synthase

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