Identification of a Residue Affecting Fatty Alcohol Selectivity in Wax Ester Synthase

Barney, Brett M.; Mann, Rachel L.; Ohlert, Janet M.

doi:10.1128/aem.02523-12

Cited by 27 publications

(68 citation statements)

References 21 publications

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“…However, actinobacterial WS/DGAT paralogs are dispersed along the bacterial clusters (53). It was recently found that there are differences in the substrate selectivities of two different R. jostii WS/DGAT enzymes (50). The finding of these different paralogs in actinobacteria could be due to differences in enzyme selectivity.…”

Section: Discussionmentioning

confidence: 89%

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Use of Limited Proteolysis and Mutagenesis To Identify Folding Domains and Sequence Motifs Critical for Wax Ester Synthase/Acyl Coenzyme A:Diacylglycerol Acyltransferase Activity

Villa

Cabezas

Cruz

et al. 2014

Appl Environ Microbiol

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Section: Discussionmentioning

confidence: 89%

“…Four hundred fifty-eight residues (97%) of Ma2 could be modeled at Ͼ90% accuracy, although the sequence identity between Ma2 and 2JPG is only 11%. The structural homology between Ma1 and 1Q9J was also recently reported (50).…”

Section: Structural Modeling Of Ws/dgat Proteins Predicts the Coadepementioning

confidence: 98%

Use of Limited Proteolysis and Mutagenesis To Identify Folding Domains and Sequence Motifs Critical for Wax Ester Synthase/Acyl Coenzyme A:Diacylglycerol Acyltransferase Activity

Villa

Cabezas

Cruz

et al. 2014

Appl Environ Microbiol

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“…Another very interesting finding concerning Ma1 from M. aquaeolei was the identification of the small alanine residue at position 360 to be relevant for the fatty alcohol chain length selectivity of Ma1 (89). The replacement of alanine by isoleucine resulted in an increased activity with the shorter-chain-length fatty alcohols nonanol and decanol compared to that of the wild-type enzyme, which is most active with dodecanol.…”

Section: Other Atfa-like Acyltransferases In Different Bacteriamentioning

confidence: 99%

“…Apart from that, the ratio of enzyme activities with hexadecanol versus dodecanol remained uninfluenced by the mutations. Therefore, the authors assumed that the incorporation of a larger amino acid residue does not block the access for longer-chain-length alcohols but improves the binding of shorter alcohols to the active site instead (89). The identification of residues affecting the substrate specificities of AtfA-like WS/DGATs is of great relevance for potential biotechnological utilization; hence, this recent study marks a first and important step in this direction.…”

Section: Other Atfa-like Acyltransferases In Different Bacteriamentioning

confidence: 99%

Acyltransferases in Bacteria

Röttig

Steinbüchel

2013

Microbiol Mol Biol Rev

148

141

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SUMMARY Long-chain-length hydrophobic acyl residues play a vital role in a multitude of essential biological structures and processes. They build the inner hydrophobic layers of biological membranes, are converted to intracellular storage compounds, and are used to modify protein properties or function as membrane anchors, to name only a few functions. Acyl thioesters are transferred by acyltransferases or transacylases to a variety of different substrates or are polymerized to lipophilic storage compounds. Lipases represent another important enzyme class dealing with fatty acyl chains; however, they cannot be regarded as acyltransferases in the strict sense. This review provides a detailed survey of the wide spectrum of bacterial acyltransferases and compares different enzyme families in regard to their catalytic mechanisms. On the basis of their studied or assumed mechanisms, most of the acyl-transferring enzymes can be divided into two groups. The majority of enzymes discussed in this review employ a conserved acyltransferase motif with an invariant histidine residue, followed by an acidic amino acid residue, and their catalytic mechanism is characterized by a noncovalent transition state. In contrast to that, lipases rely on completely different mechanism which employs a catalytic triad and functions via the formation of covalent intermediates. This is, for example, similar to the mechanism which has been suggested for polyester synthases. Consequently, although the presented enzyme types neither share homology nor have a common three-dimensional structure, and although they deal with greatly varying molecule structures, this variety is not reflected in their mechanisms, all of which rely on a catalytically active histidine residue.

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“…2.6‐ or 1.8‐fold higher when compared to the wild‐type AtfA, respectively. Ma1(A360I) showed a significant decrease in its activity toward n‐hexadecanol and n‐dodecanol, whereas the activity with nonanol was even 5.4‐fold higher when compared to the activity of the non‐mutated Ma1 enzyme . Although activities toward alcohols shorter than nonanol were not described in the latter publication, both enzyme variants seemed to be interesting enzymes for our study.…”

Section: Introductionmentioning

confidence: 76%

In vitro characterization of five bacterial WS/DGAT acyltransferases regarding the synthesis of biotechnologically relevant short‐chain‐length esters

Röttig

Wolf

Steinbüchel

2015

Euro J Lipid Sci & Tech

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For the biotechnical production of biofuels, oleo‐, or fine chemicals bacterial wax ester synthase/acyl‐Coenzyme A:diacylglycerol acyltransferases (WS/DGAT) are discussed as interesting candidates for in vivo esterification reactions. In this study, the suitability of selected acyltransferases for the conversion of non‐physiological substrates like short‐chain‐length alcohols and short‐chain length or even branched acyl‐CoAs has been investigated. In vitro analyzes of purified AtfA and AtfA(G355I) from Acinetobacter baylyi, Ma1(A360I) from Marinobacter aquaeolei, WS2 from Marinobacter hydrocarbonoclasticus, and AtfA1 from Alcanivorax borkumensis were conducted to evaluate the specific activities of these enzymes toward n‐hexadecanol (C16), n‐dodecanol (C12), ethanol (C2), and methanol (C1), palmitoyl‐CoA (C16), butyryl‐CoA (C4) as well as toward branched 3‐hydroxybutyryl‐CoA and 2‐hydroxyisobutyryl‐CoA. Athough long‐ and medium‐chain‐length substrates were preferred by all five enzymes, WS2 and AtfA showed the highest relative activities with ethanol or methanol when compared to n‐hexadecanol, whereas residual activities toward short or branched acyl‐CoAs could only be measured with AtfA and AtfA1. Practical applications: Bacterial WS/DGATs can be used for in vivo and in vitro approaches to synthesize custom‐made lipids, such as triglycerides or wax esters. However, due to their broad substrate ranges these enzymes are also promising candidates for the synthesis of other, industrially valuable oleo‐ and fine chemicals. Short‐chain‐length esters are important intermediates and building blocks for many production processes and, at present, there is a great demand for enzymes which are able to catalyze their synthesis. The physiological substrates of bacterial WS/DGAT enzymes are medium‐ to long‐chain length acyl‐CoAs and fatty alcohols to synthesize medium‐ to long‐chain length wax esters. In this study, we investigated five different bacterial WS/DGATs for their ability to synthesize short‐chain length esters, which represent biotechnologically interesting compounds. The enzymes AtfA, AtfA(G355I), Ma1(A360I), WS2, and AtfA1 were selected, purified, and characterized in vitro. In addition to the reference substrates, hexadecanol and palmitoyl‐CoA, their activity with dodecanol, ethanol, or methanol, on the one hand, and lauryl‐CoA, butyryl‐CoA, 3‐hydroxybutyryl‐CoA, or 2‐hydroxyisobutyryl‐CoA, on the other hand, were studied and compared.

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Identification of a Residue Affecting Fatty Alcohol Selectivity in Wax Ester Synthase

Cited by 27 publications

References 21 publications

Use of Limited Proteolysis and Mutagenesis To Identify Folding Domains and Sequence Motifs Critical for Wax Ester Synthase/Acyl Coenzyme A:Diacylglycerol Acyltransferase Activity

Use of Limited Proteolysis and Mutagenesis To Identify Folding Domains and Sequence Motifs Critical for Wax Ester Synthase/Acyl Coenzyme A:Diacylglycerol Acyltransferase Activity

Acyltransferases in Bacteria

In vitro characterization of five bacterial WS/DGAT acyltransferases regarding the synthesis of biotechnologically relevant short‐chain‐length esters

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