Three-dimensional structures of enzymes useful for ?-lactam antibiotic production

Barends, T.R.M.; Yoshida, Hiromi; Dijkstra, Bauke W.

doi:10.1016/j.copbio.2004.06.009

Cited by 18 publications

(12 citation statements)

References 26 publications

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“…Deacetyl cephalosporins are valuable building blocks for the production of semisynthetic β‐lactam antibiotics. These compounds are derived from cephalosporin C or 7‐ACA via enzymatic or chemical processes 10. The thermostable TM0077 esterase may be valuable in the preparation of derivatives of β‐lactam antibiotics.…”

Section: Resultsmentioning

confidence: 99%

“…Cephalosporins belong to the β‐lactam class of antibiotics, which also includes penicillin, and affect bacterial cell growth by inhibiting the penicillin binding protein that cross‐links peptide glycans required for cell wall formation 9. The production of deacetylated cephalosporins is of great interest because these compounds are valuable building blocks for the production of semi‐synthetic β‐lactam antibiotics 10, 11…”

Section: Introductionmentioning

confidence: 99%

“…9 The production of deacetylated cephalosporins is of great interest because these compounds are valuable building blocks for the production of semi-synthetic b-lactam antibiotics. 10,11 To explore the catalytic capacity of the predicted AXE from T. maritima and gain a better insight into the structure and function of the family 7 carbohydrate esterases, TM0077 was expressed and purified, and three-dimensional structures of the native enzyme and its complexes with phenylmethylsulfonyl fluoride (PMSF) and paraoxon inhibitors were determined by X-ray crystallography. Furthermore, the enzyme was functionally characterized, and various biochemical properties including the positional specificity of the esterase were investigated.…”

Section: Introductionmentioning

confidence: 99%

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Functional and structural characterization of a thermostable acetyl esterase from Thermotoga maritima

et al. 2012

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TM0077 from Thermotoga maritima is a member of the carbohydrate esterase family 7 and is active on a variety of acetylated compounds, including cephalosporin C. TM0077 esterase activity is confined to short-chain acyl esters (C2-C3), and is optimal around 100°C and pH 7.5. The positional specificity of TM0077 was investigated using 4-nitrophenyl-β-D-xylopyranoside monoacetates as substrates in a β-xylosidase-coupled assay. TM0077 hydrolyzes acetate at positions 2, 3 and 4 with equal efficiency. No activity was detected on xylan or acetylated xylan, which implies that TM0077 is an acetyl esterase and not an acetyl xylan esterase as currently annotated. Selenomethionine-substituted and native structures of TM0077 were determined at 2.1 Å and 2.5 Å resolution, respectively, revealing a classic α/β-hydrolase fold. TM0077 assembles into a doughnut-shaped hexamer with small tunnels on either side leading to an inner cavity, which contains the six catalytic centers. Structures of TM0077 with covalently bound phenylmethylsulfonyl fluoride (PMSF) and paraoxon were determined to 2.4 Å and 2.1 Å, respectively, and confirmed that both inhibitors bind covalently to the catalytic serine (Ser188). Upon binding of inhibitor, the catalytic serine adopts an altered conformation, as observed in other esterase and lipases, and supports a previously proposed catalytic mechanism in which this Ser hydroxyl rotation prevents reversal of the reaction and allows access of a water molecule for completion of the reaction.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Functional and structural characterization of a thermostable acetyl esterase from Thermotoga maritima

et al. 2012

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“…This enzyme has the same fold as DmpA, but catalyzes the reversible transfer of an acetyl group from N-acetylornithine to glutamate (Elkins et al, 2005). Among the Ntn hydrolases, penicillin and cephalosporin acylases as well as b-aminopeptidases have particularly high potential for biocatalytic conversions (Bruggink et al, 1998;Barends et al, 2004;Heck et al, 2007Heck et al, , 2009Heck et al, , 2010.…”

Section: Introductionmentioning

confidence: 99%

Autoproteolytic and Catalytic Mechanisms for the β-Aminopeptidase BapA—A Member of the Ntn Hydrolase Family

et al. 2012

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The β-aminopeptidase BapA from Sphingosinicella xenopeptidilytica belongs to the N-terminal nucleophile (Ntn) hydrolases of the DmpA-like family and has the unprecedented property of cleaving N-terminal β-amino acid residues from peptides. We determined the crystal structures of the native (αβ)₄ heterooctamer and of the 153 kDa precursor homotetramer at a resolution of 1.45 and 1.8 Å, respectively. These structures together with mutational analyses strongly support mechanisms for autoproteolysis and catalysis that involve residues Ser250, Ser288, and Glu290. The autoproteolytic mechanism is different from the one so far described for Ntn hydrolases. The structures together with functional data also provide insight into the discriminating features of the active site cleft that determine substrate specificity.

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“…This in vivo process is exploited for the commercial production of penicillins [1,2]. Starting from penicillin G, obtained from fermentation of P. chrysogenum, several other -lactam antibiotics can be made via semisynthesis [3,4]. In this production process, the side chain is Wrst removed from the -lactam core, after which the -lactam nucleus is acylated with the side chain of interest in an enzymatic or non-enzymatic process.…”

mentioning

confidence: 99%

An approach to prevent aggregation during the purification and crystallization of wild type acyl coenzyme A: Isopenicillin N acyltransferase from Penicillium chrysogenum

Yoshida

Hensgens²,

Laan

et al. 2005

Protein Expression and Purification

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Three-dimensional structures of enzymes useful for ?-lactam antibiotic production

Cited by 18 publications

References 26 publications

Functional and structural characterization of a thermostable acetyl esterase from Thermotoga maritima

Functional and structural characterization of a thermostable acetyl esterase from Thermotoga maritima

Autoproteolytic and Catalytic Mechanisms for the β-Aminopeptidase BapA—A Member of the Ntn Hydrolase Family

An approach to prevent aggregation during the purification and crystallization of wild type acyl coenzyme A: Isopenicillin N acyltransferase from Penicillium chrysogenum

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