Verstrepen Kevin J, Van Laere Stijn D M, Vanderhaegen Bart M P, Derdelinckx Guy, Dufour Jean-Pierre, Pretorius Isak S, Winderickx Joris, Thevelein Johan M, Delvaux Freddy R
Centre for Malting and Brewing Science, Department of Food and Microbial Technology, Katholieke Universiteit Leuven, B-3001 Louvain (Heverlee), Belgium.
Appl Environ Microbiol. 2003 Sep;69(9):5228-37. doi: 10.1128/AEM.69.9.5228-5237.2003.
Volatile aroma-active esters are responsible for the fruity character of fermented alcoholic beverages such as beer and wine. Esters are produced by fermenting yeast cells in an enzyme-catalyzed intracellular reaction. In order to investigate and compare the roles of the known Saccharomyces cerevisiae alcohol acetyltransferases, Atf1p, Atf2p and Lg-Atf1p, in volatile ester production, the respective genes were either deleted or overexpressed in a laboratory strain and a commercial brewing strain. Subsequently, the ester formation of the transformants was monitored by headspace gas chromatography and gas chromatography combined with mass spectroscopy (GC-MS). Analysis of the fermentation products confirmed that the expression levels of ATF1 and ATF2 greatly affect the production of ethyl acetate and isoamyl acetate. GC-MS analysis revealed that Atf1p and Atf2p are also responsible for the formation of a broad range of less volatile esters, such as propyl acetate, isobutyl acetate, pentyl acetate, hexyl acetate, heptyl acetate, octyl acetate, and phenyl ethyl acetate. With respect to the esters analyzed in this study, Atf2p seemed to play only a minor role compared to Atf1p. The atf1Delta atf2Delta double deletion strain did not form any isoamyl acetate, showing that together, Atf1p and Atf2p are responsible for the total cellular isoamyl alcohol acetyltransferase activity. However, the double deletion strain still produced considerable amounts of certain other esters, such as ethyl acetate (50% of the wild-type strain), propyl acetate (50%), and isobutyl acetate (40%), which provides evidence for the existence of additional, as-yet-unknown ester synthases in the yeast proteome. Interestingly, overexpression of different alleles of ATF1 and ATF2 led to different ester production rates, indicating that differences in the aroma profiles of yeast strains may be partially due to mutations in their ATF genes.
挥发性香气活性酯赋予了啤酒和葡萄酒等发酵酒精饮料的果香味。酯类是由发酵酵母细胞在酶催化的细胞内反应中产生的。为了研究和比较已知的酿酒酵母酒精乙酰转移酶Atf1p、Atf2p和Lg-Atf1p在挥发性酯类产生中的作用，分别在一个实验室菌株和一个商业酿造菌株中对各自的基因进行了缺失或过表达操作。随后，通过顶空气相色谱法以及气相色谱 - 质谱联用（GC-MS）对转化体的酯形成情况进行监测。对发酵产物的分析证实，ATF1和ATF2的表达水平极大地影响乙酸乙酯和乙酸异戊酯的产生。GC-MS分析表明，Atf1p和Atf2p还负责形成多种挥发性较低的酯类，如乙酸丙酯、乙酸异丁酯、乙酸戊酯、乙酸己酯、乙酸庚酯、乙酸辛酯和苯乙酸乙酯。就本研究中分析的酯类而言，与Atf1p相比，Atf2p似乎只起次要作用。atf1Delta atf2Delta双缺失菌株不形成任何乙酸异戊酯，这表明Atf1p和Atf2p共同负责细胞总的异戊醇乙酰转移酶活性。然而，双缺失菌株仍产生相当数量的某些其他酯类，如乙酸乙酯（野生型菌株的50%）、乙酸丙酯（50%）和乙酸异丁酯（40%），这为酵母蛋白质组中存在其他尚未知晓的酯合成酶提供了证据。有趣的是，ATF1和ATF2不同等位基因的过表达导致了不同的酯产生速率，这表明酵母菌株香气特征的差异可能部分归因于其ATF基因的突变。
