Short-chain esters contribute to the blend of volatiles that define the strawberry aroma. The last step in their biosynthesis involves an alcohol acyltransferase that catalyses the esterification of an acyl moiety of acyl-CoA with an alcohol. This study identified a novel strawberry alcohol acyltransferase gene (FaAAT2) whose expression pattern during fruit receptacle growth and ripening is in accordance with the production of esters throughout strawberry fruit ripening. The full-length FaAAT2 cDNA was cloned and expressed in Escherichia coli and its activity was analysed with acyl-CoA and alcohol substrates. The semi-purified FaAAT2 enzyme had activity with C1-C8 straight-chain alcohols and aromatic alcohols in the presence of acetyl-CoA. Cinnamyl alcohol was the most efficient acyl acceptor. When FaAAT2 expression was transiently downregulated in the fruit receptacle by agroinfiltration, the volatile ester production was significantly reduced in strawberry fruit. The results suggest that FaAAT2 plays a significant role in the production of esters that contribute to the final strawberry fruit flavour.

Background: Fragaria x ananassa enone oxidoreductase catalyzes the ripening-induced formation of 4-hydroxy-2,5-dimethyl-3(2H)-furanone in strawberries. Results: By determining six x-ray structures of different substrate complexes the enzymatic mechanism was elucidated and experimentally confirmed by deuterium labeling. Conclusion: The 4R-hydride of NAD(P)H is transferred to an exo-cyclic carbon double bond. Significance: Enzymatic 4-hydroxy-2,5-dimethyl-3(2H)-furanone synthesis reveals a new reaction mechanism and advances understanding of a biotechnologically relevant biosynthetic pathway.

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bEnzymatic transamination of amino acids yields ␣-keto acids and is the initial step for the production of volatile compounds that contribute to the sensory perception of fermented foods such as salami. Lactobacillus sakei is one of the lactic acid bacterial strains commonly used in starter cultures. Although the genome sequence of L. sakei 23K lacks genes encoding typical branchedchain amino acid transaminases, transamination activity and the formation of amino acid-derived volatile metabolites could be demonstrated. A protein purified from L. sakei is held responsible for the transamination activity. By heterologous expression of the corresponding gene in Escherichia coli, we were able to characterize the transamination side activity of an enzyme annotated as a putative acylphosphatase (AcP). A transamination side activity of hen egg white lysozyme (HEWL) was also discovered. Both enzymes showed substrate specificity toward branched-chain and aromatic amino acids. AcP also accepted L-methionine. Activity was optimal at neutral pH for both enzymes, whereas AcP showed a significantly higher temperature optimum (55°C) than that of HEWL (37°C). Kinetic parameters revealed high affinity toward L-leucine for AcP (K m ‫؍‬ 1.85 mM) and toward L-isoleucine for HEWL (K m ‫؍‬ 3.79 mM). AcP seems to play a major role in the metabolism of amino acids in L. sakei. The flavor of dry fermented sausages derives from the ingredients (meat, spices, and smoke) and the chemical changes occurring during the fermentation and drying process. Flavor formation in dry fermented sausage occurs by meat enzymes and bacterial fermentation, which are responsible for the metabolism of fats, carbohydrates, and proteins (1, 2). The autochthonous microbiota of fermented meat products is dominated by coagulase-negative staphylococci and lactic acid bacteria (LAB), mainly Lactobacillus sakei, Lactobacillus curvatus, and Lactobacillus plantarum (3, 4). L. sakei, formerly known as L. sake, is one of the dominating LAB in dry spontaneously fermented sausages (3, 5). Therefore, it is commonly used in starter cultures for the production of dry fermented sausages (6).Free amino acids, incorporated into LAB cells by amino acid transporters or released intracellularly by peptidase activity, are converted, among other metabolic pathways, to various volatile compounds by transamination and decarboxylation. Enzymatic transamination leading to the formation of the corresponding ␣-keto acids is the first step of this intracellular amino acid catabolism to volatile metabolites in LAB (Fig. 1) (2, 7). The transformation of branched-chain, aromatic, and sulfur-containing amino acids by these reactions yields volatile compounds, which contribute to the sensory perception of fermented food (8, 9). It has been calculated that 11.8% of the volatile compounds in salami probably originate from amino acid catabolism (10).Analysis of the genome sequence of L. sakei 23K delivered insights into the strain's potential concerning aroma formation from amino acid substrates. G...