Crystal Structure of Homoserine Transacetylase from <i>Haemophilus influenzae</i> Reveals a New Family of α/β-Hydrolases<sup>,</sup>

Mirza, I.A.; Nazi, Ishac; Korczynska, Magdalena; Wright, Gerard D.; Berghuis, Albert M.

doi:10.1021/bi051951y

Cited by 48 publications

(59 citation statements)

References 25 publications

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“…A promising target in this pathway is HTA, which catalyzes the first committed step in the biosynthesis of Met from Asp. HTA is present in several pathogenic bacterial species such as H. influenzae, 7,18,30 Pseudomonas aeruginosa, Mycobacterium tuberculosis, 31 Leptospira meyeri 32 and most fungi. 8,9,33 HTA uses a classic Ser/His/Asp catalytic triad to promote transfer of the acetyl group from acetyl-CoA to homoserine.…”

Section: Discussionmentioning

confidence: 99%

“…7 The construct was introduced into Escherichia coli BL21 (DE3) allowing for the expression of HTA Hin with an N-terminal hexa-histidine tag.…”

Section: Hta Expression and Purificationmentioning

confidence: 99%

“…5 The absence of this pathway in mammals makes it an attractive target for antimicrobial drug discovery. Small molecule inhibitors have been discovered and used as probes of the catalytic and biochemical functions of the different enzymes of this pathway including cystathionine b-lyase, 6 homoserine transacetylase (HTA), [7][8][9][10][11] Met synthase (MET6) 2 and homoserine dehydrogenase (HOM6). 3,[12][13][14] Moreover, gene disruption methods have identified the importance of these enzymes in pathogenesis in several animal models of virulence.…”

Section: Introductionmentioning

confidence: 99%

“…5 Extensive biochemical and structural studies have been performed on HTA from the fission yeast Schizosaccharomyces pombe 9 and the Gramnegative bacterial pathogen Haemophilus influenzae. 7,18 Genetic studies in bacteria and yeast have shown that deletion of the gene that encodes for HTA is lethal in minimal media and Met supplementation is required for cell viability. 8,9,[19][20][21] Furthermore, the gene encoding HTA, MET2, has been found to be required for virulence in the human pathogen Cryptococcus neoformans.…”

Section: Introductionmentioning

confidence: 99%

“…The three-dimensional structure of HTA reveals an elongated substrate-binding tunnel that leads to the nucleophile Ser in the active site involved in catalysis ( Figure 2). 7 Potential inactivators that take advantage of both modification of the active site Ser and the substrate-binding tunnel should be good probes of HTA function and potential leads for antimicrobial agents. We describe the inactivation of H. influenzae HTA by natural product and synthetic b-lactones and use these to biochemically confirm that Ser143 is the active site nucleophile and evaluate their antibiotic activity.…”

Section: Introductionmentioning

confidence: 99%

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β-Lactone natural products and derivatives inactivate homoserine transacetylase, a target for antimicrobial agents

et al. 2011

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Homoserine transacetylase (HTA) catalyzes the transfer of an acetyl group from acetyl-CoA to the hydroxyl group of homoserine. This is the first committed step in the biosynthesis of methionine (Met) from aspartic acid in many fungi, Gram-positive and some Gram-negative bacteria. The enzyme is absent in higher eukaryotes and is important for microorganism growth in Met-poor environments, such as blood serum, making HTA an attractive target for new antimicrobial agents. HTA catalyzes acetyl transfer via a double displacement mechanism facilitated by a classic Ser-His-Asp catalytic triad located at the bottom of a narrow actives site tunnel. We explored the inhibitory activity of several b-lactones to block the activity of HTA. In particular, the natural product ebelactone A, a b-lactone with a hydrophobic tail was found to be a potent inactivator of HTA from Haemophilus influenzae. Synthetic analogs of ebelactone A demonstrated improved inactivation characteristics. Covalent modification of HTA was confirmed by mass spectrometry, and peptide mapping identified Ser143 as the modified residue, consistent with the known structure and mechanism of the enzyme. These results demonstrate that b-lactone inhibitors are excellent biochemical probes of HTA and potential leads for new antimicrobial agents.

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Section: Discussionmentioning

confidence: 99%

“…7 The construct was introduced into Escherichia coli BL21 (DE3) allowing for the expression of HTA Hin with an N-terminal hexa-histidine tag.…”

Section: Hta Expression and Purificationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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β-Lactone natural products and derivatives inactivate homoserine transacetylase, a target for antimicrobial agents

et al. 2011

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A novel esterase subfamily with α/β‐hydrolase fold suggested by structures of two bacterial enzymes homologous to l‐homoserine O‐acetyl transferases

et al. 2015

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MekB from Pseudomonas veronii and CgHle from Corynebacteriumglutamicum belong to the superfamily of α/β‐hydrolase fold proteins. Based on sequence comparisons, they are annotated as homoserine transacetylases in popular databases like UNIPROT, PFAM or ESTHER. However, experimentally, MekB and CgHle were shown to be esterases that hydrolyse preferentially acetic acid esters. We describe the x‐ray structures of these enzymes solved to high resolution. The overall structures confirm the close relatedness to experimentally validated homoserine acetyl transferases, but simultaneously the structures exclude the ability of MekB and CgHle to bind homoserine and acetyl‐CoA. Insofar the MekB and CgHle structures suggest dividing the homoserine transacetylase family into subfamilies, namely genuine acetyl transferases and acetyl esterases with MekB and CgHle as constituting members of the latter.

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Amino Acid Biosynthesis

Parker

Pratt

2010

Amino Acids, Peptides and Proteins in Organic Chemistry

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The ribosomal synthesis of proteins utilizes a family of 20 a-amino acids that are universally coded by the translation machinery; in addition, two further a-amino acids, selenocysteine and pyrrolysine, are now believed to be incorporated into proteins via ribosomal synthesis in some organisms. More than 300 other amino acid residues have been identified in proteins, but most are of restricted distribution and produced via post-translational modification of the ubiquitous protein amino acids [1]. The ribosomally encoded a-amino acids described here ultimately derive from a-keto acids by a process corresponding to reductive amination. The most important biosynthetic distinction relates to whether appropriate carbon skeletons are pre-existing in basic metabolism or whether they have to be synthesized de novo and this division underpins the structure of this chapter.There are a small number of a-keto acids ubiquitously found in core metabolism, notably pyruvate (and a related 3-phosphoglycerate derivative from glycolysis), together with two components of the tricarboxylic acid cycle (TCA), oxaloacetate and a-ketoglutarate (a-KG). These building blocks ultimately provide the carbon skeletons for unbranched a-amino acids of three, four, and five carbons, respectively. a-Amino acids with shorter (glycine) or longer (lysine and pyrrolysine) straight chains are made by alternative pathways depending on the available raw materials. The strategic challenge for the biosynthesis of most straight-chain amino acids centers around two issues: how is the a-amino function introduced into the carbon skeleton and what functional group manipulations are required to generate the diversity of side-chain functionality required for the protein function?The core family of straight-chain amino acids does not provide all the functionality required for proteins. a-Amino acids with branched side-chains are used for two purposes; the primary need is related to protein structural issues. Proteins fold into well-defined three-dimensional shapes by virtue of their amphipathic nature: a significant fraction of the amino acid side-chains are of low polarity and the hydrophobic effect drives the formation of ordered structures in which these side-chains are buried away from water. In contrast to the straight-chain amino

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Crystal Structure of Homoserine Transacetylase from Haemophilus influenzae Reveals a New Family of α/β-Hydrolases^,

Cited by 48 publications

References 25 publications

β-Lactone natural products and derivatives inactivate homoserine transacetylase, a target for antimicrobial agents

β-Lactone natural products and derivatives inactivate homoserine transacetylase, a target for antimicrobial agents

A novel esterase subfamily with α/β‐hydrolase fold suggested by structures of two bacterial enzymes homologous to l‐homoserine O‐acetyl transferases

Amino Acid Biosynthesis

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Crystal Structure of Homoserine Transacetylase from Haemophilus influenzae Reveals a New Family of α/β-Hydrolases,

Cited by 48 publications

References 25 publications

β-Lactone natural products and derivatives inactivate homoserine transacetylase, a target for antimicrobial agents

β-Lactone natural products and derivatives inactivate homoserine transacetylase, a target for antimicrobial agents

A novel esterase subfamily with α/β‐hydrolase fold suggested by structures of two bacterial enzymes homologous to l‐homoserine O‐acetyl transferases

Amino Acid Biosynthesis

Contact Info

Product

Resources

About

Crystal Structure of Homoserine Transacetylase from Haemophilus influenzae Reveals a New Family of α/β-Hydrolases^,