Garbe Leif-Alexander, Barbosa de Almeida Roberto, Nagel Renate, Wackerbauer Karl, Tressl Roland
Institut für Biotechnologie, Molekularanalytik, Technische Universität Berlin, Seestrasse 13, D-13353 Berlin, Germany.
J Agric Food Chem. 2006 Feb 8;54(3):946-55. doi: 10.1021/jf051993t.
The lipoxygenase isoenzymes LOX1 and LOX2 from green malt were separated by isoelectric focusing, and their catalytic properties regarding complex lipids as substrates were characterized. The regio- and stereoisomers of hydroperoxy octadecadienoates (HPODE) resulting from LOX1 and LOX2 enzymatic transformations of linoleic acid, methyl linoleate, linoleic acid glycerol esters monolinolein, dilinolein, and trilinolein, and 1-palmitoyl-2-linoleoyl-glycero-3-phosphocholine (PamLinGroPCho) were determined. In addition, biotransformations of polar and nonpolar lipids extracted from malt were performed with LOX1 and LOX2. The results show that LOX2 catalyzes the oxidation of esterified fatty acids at a higher rate and is more regioselective than LOX1. The dual position specificity of LOX2 (9-HPODE:13-HPODE) with trilinolein as the substrate (6:94) was higher than the resultant ratio (13:87) when free linoleic acid was transformed. A high (S)-enantiomeric excess of 13-HPODE was analyzed with all esterified substrates confirming the formation of 13-HPODE through the LOX2 enzyme; however, 9-HPODE detected after LOX2 biotransformations showed (R)-enantiomeric excesses. PamLinGroPCho was oxygenated by LOX1 with the highest regio- and stereoselectivities among the applied substrates.
通过等电聚焦分离了绿色麦芽中的脂氧合酶同工酶LOX1和LOX2，并对它们以复合脂质为底物的催化特性进行了表征。测定了由亚油酸、亚油酸甲酯、亚油酸甘油酯单亚油酸甘油酯、二亚油酸甘油酯、三亚油酸甘油酯以及1-棕榈酰-2-亚油酰-甘油-3-磷酸胆碱（PamLinGroPCho）经LOX1和LOX2酶促转化产生的氢过氧化十八碳二烯酸（HPODE）的区域异构体和立体异构体。此外，用LOX1和LOX2对从麦芽中提取的极性和非极性脂质进行了生物转化。结果表明，LOX2催化酯化脂肪酸氧化的速率更高，且比LOX1具有更高的区域选择性。以三亚油酸甘油酯为底物时，LOX2的双重位置特异性（9-HPODE:13-HPODE）为6:94，高于转化游离亚油酸时的比例（13:87）。对所有酯化底物分析发现，13-HPODE具有较高的（S）-对映体过量，证实了通过LOX2酶形成了13-HPODE；然而，LOX2生物转化后检测到的9-HPODE显示出（R）-对映体过量。在所应用的底物中，PamLinGroPCho被LOX1氧化时具有最高的区域和立体选择性。
