Homoserine acetyltransferase (homoserine transacetylase; EC: 2.3.1.31) catalyses the first step unique to methionine biosynthesis, converting L-homoserine to O-acetyl-L-homoserine using acetyl-CoA as an acetyl group donor. This enzyme is unique to fungi and some bacteria. This enzyme regulates homoserine in a number of biosynthetic pathways, making it vital to cell growth and viability. The biosynthesis of methionine in particular has been shown to be susceptible to small-molecule inhibition in fungi and is consequently a good candidate for antifungal agents PIRSF000443. A subset of this family was shown by Toelzer et al. to be esterases and unable to bind acetyl-CoA and was extracted as a new family (Homoserine_transacetylase_like_est)
Homoserine transacetylase is a required catalyst in the biochemical pathway that metabolizes Asp to Met in fungi. The enzyme from the yeast Schizosaccharomyces pombe activates the hydroxyl group of L-homoserine by acetylation from acetyl coenzyme A. This enzyme is unique to fungi and some bacteria and presents an important new target for drug discovery. Steady-state kinetic parameters provide evidence that this enzyme follows a ping-pong mechanism. Proton inventory was consistent with a single-proton transfer, and pH studies suggested the participation of at least one residue with a pKa value of 6.4-6.6, possibly a His or Asp/Glu in catalysis. Protein sequence alignments indicate that this enzyme belongs to the alpha/beta-hydrolase fold superfamily of enzymes, indicating the involvement of an active-site nucleophile and possibly a canonical catalytic triad. We constructed site-specific mutants and identified Ser163, Asp403, and His432 as the likely active-site residues of a catalytic triad based on steady-state kinetics and genetic complementation of a yeast null mutant. Moreover, unlike the wild-type enzyme, inactive site mutants were not capable of producing an acetyl-enzyme intermediate. Homoserine transacetylase therefore catalyzes the acetylation of L-homoserine via a covalent acyl-enzyme intermediate through an active-site Ser. These results form the basis of future exploitation of this enzyme as an antimicrobial target.
        
Title: Enzyme-catalyzed acylation of homoserine: mechanistic characterization of the Haemophilus influenzae met2-encoded homoserine transacetylase Born TL, Franklin M, Blanchard JS Ref: Biochemistry, 39:8556, 2000 : PubMed
The first unique step in bacterial and plant methionine biosynthesis involves the acylation of the gamma-hydroxyl of homoserine. In Haemophilus influenzae, acylation is accomplished via an acetyl-CoA-dependent acetylation catalyzed by homoserine transacetylase. The activity of this enzyme regulates flux of homoserine into multiple biosynthetic pathways and, therefore, represents a critical control point for cell growth and viability. We have cloned homoserine transacetylase from H. influenzae and present the first detailed enzymatic study of this enzyme. Steady-state kinetic experiments demonstrate that the enzyme utilizes a ping-pong kinetic mechanism in which the acetyl group of acetyl-CoA is initially transferred to an enzyme nucleophile before subsequent transfer to homoserine to form the final product, O-acetylhomoserine. The maximal velocity and V/K(homoserine) were independent of pH over the range of values tested, while V/K(acetyl)(-)(CoA) was dependent upon the ionization state of a single group exhibiting a pK value of 8.6, which was required to be protonated. Solvent kinetic isotope effect studies yielded inverse effects of 0.75 on V and 0.74 on V/K(CoA) on the reverse reaction and effects of 1.2 on V and 1.7 on V/K(homoserine) on the forward reaction. Direct evidence for the formation of an acetyl-enzyme intermediate was obtained using rapid-quench labeling studies. On the basis of these observations, we propose a chemical mechanism for this important member of the acyltransferase family and contrast its mechanism with that of homoserine transsuccinylase.
A 5.1-kb DNA fragment from Saccharomyces cerevisiae, which complements a yeast met2 mutant strain, has been cloned. This fragment contains the wild-type MET2 gene which codes for the homoserine O-transacetylase, one of the methionine biosynthetic enzymes. The presence of the MET2 gene has been shown by integrative transformation experiments and genetic analyses of the resulting transformants. The complete nucleotide sequence of a 2826-bp DNA fragment carrying the MET2 gene has been determined. The sequence contains one major open reading frame of 438 codons, giving a calculated Mr of 48,370 for the encoded protein. We have identified the transcriptional product of the MET2 gene and estimated its size at 1650 nucleotides.
Homoserine transacetylase catalyzes one of the required steps in the biosynthesis of methionine in fungi and several bacteria. We have determined the crystal structure of homoserine transacetylase from Haemophilus influenzae to a resolution of 1.65 A. The structure identifies this enzyme to be a member of the alpha/beta-hydrolase structural superfamily. The active site of the enzyme is located near the end of a deep tunnel formed by the juxtaposition of two domains and incorporates a catalytic triad involving Ser143, His337, and Asp304. A structural basis is given for the observed double displacement kinetic mechanism of homoserine transacetylase. Furthermore, the properties of the tunnel provide a rationale for how homoserine transacetylase catalyzes a transferase reaction vs hydrolysis, despite extensive similarity in active site architecture to hydrolytic enzymes.
        
Title: Catalytic mechanism of fungal homoserine transacetylase Nazi I, Wright GD Ref: Biochemistry, 44:13560, 2005 : PubMed
Homoserine transacetylase is a required catalyst in the biochemical pathway that metabolizes Asp to Met in fungi. The enzyme from the yeast Schizosaccharomyces pombe activates the hydroxyl group of L-homoserine by acetylation from acetyl coenzyme A. This enzyme is unique to fungi and some bacteria and presents an important new target for drug discovery. Steady-state kinetic parameters provide evidence that this enzyme follows a ping-pong mechanism. Proton inventory was consistent with a single-proton transfer, and pH studies suggested the participation of at least one residue with a pKa value of 6.4-6.6, possibly a His or Asp/Glu in catalysis. Protein sequence alignments indicate that this enzyme belongs to the alpha/beta-hydrolase fold superfamily of enzymes, indicating the involvement of an active-site nucleophile and possibly a canonical catalytic triad. We constructed site-specific mutants and identified Ser163, Asp403, and His432 as the likely active-site residues of a catalytic triad based on steady-state kinetics and genetic complementation of a yeast null mutant. Moreover, unlike the wild-type enzyme, inactive site mutants were not capable of producing an acetyl-enzyme intermediate. Homoserine transacetylase therefore catalyzes the acetylation of L-homoserine via a covalent acyl-enzyme intermediate through an active-site Ser. These results form the basis of future exploitation of this enzyme as an antimicrobial target.
        
Title: Enzyme-catalyzed acylation of homoserine: mechanistic characterization of the Haemophilus influenzae met2-encoded homoserine transacetylase Born TL, Franklin M, Blanchard JS Ref: Biochemistry, 39:8556, 2000 : PubMed
The first unique step in bacterial and plant methionine biosynthesis involves the acylation of the gamma-hydroxyl of homoserine. In Haemophilus influenzae, acylation is accomplished via an acetyl-CoA-dependent acetylation catalyzed by homoserine transacetylase. The activity of this enzyme regulates flux of homoserine into multiple biosynthetic pathways and, therefore, represents a critical control point for cell growth and viability. We have cloned homoserine transacetylase from H. influenzae and present the first detailed enzymatic study of this enzyme. Steady-state kinetic experiments demonstrate that the enzyme utilizes a ping-pong kinetic mechanism in which the acetyl group of acetyl-CoA is initially transferred to an enzyme nucleophile before subsequent transfer to homoserine to form the final product, O-acetylhomoserine. The maximal velocity and V/K(homoserine) were independent of pH over the range of values tested, while V/K(acetyl)(-)(CoA) was dependent upon the ionization state of a single group exhibiting a pK value of 8.6, which was required to be protonated. Solvent kinetic isotope effect studies yielded inverse effects of 0.75 on V and 0.74 on V/K(CoA) on the reverse reaction and effects of 1.2 on V and 1.7 on V/K(homoserine) on the forward reaction. Direct evidence for the formation of an acetyl-enzyme intermediate was obtained using rapid-quench labeling studies. On the basis of these observations, we propose a chemical mechanism for this important member of the acyltransferase family and contrast its mechanism with that of homoserine transsuccinylase.
A 5.1-kb DNA fragment from Saccharomyces cerevisiae, which complements a yeast met2 mutant strain, has been cloned. This fragment contains the wild-type MET2 gene which codes for the homoserine O-transacetylase, one of the methionine biosynthetic enzymes. The presence of the MET2 gene has been shown by integrative transformation experiments and genetic analyses of the resulting transformants. The complete nucleotide sequence of a 2826-bp DNA fragment carrying the MET2 gene has been determined. The sequence contains one major open reading frame of 438 codons, giving a calculated Mr of 48,370 for the encoded protein. We have identified the transcriptional product of the MET2 gene and estimated its size at 1650 nucleotides.
        
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No structure scheme yet for this family
Structures in Homoserine_transacetylase family (18)