Zheng X, Seiliez I, Hastings N, Tocher D R, Panserat S, Dickson C A, Bergot P, Teale A J
Institute of Aquaculture, University of Stirling, Airthrey Road, Stirling FK9 4LA, Scotland, United Kingdom.
Comp Biochem Physiol B Biochem Mol Biol. 2004 Oct;139(2):269-79. doi: 10.1016/j.cbpc.2004.08.003.
Fish are the most important dietary source of the n-3 highly unsaturated fatty acids (HUFA), eicosapentaenoic (EPA) and docosahexaenoic acid (DHA), that have particularly important roles in human nutrition reflecting their roles in critical physiological processes. The objective of the study described here was to clone, functionally characterize and compare expressed fatty acid desaturase genes involved in the production of EPA and DHA in freshwater and marine teleost fish species. Putative fatty acid desaturase cDNAs were isolated and cloned from common carp (Cyprinus carpio) and turbot (Psetta maximus). The enzymic activities of the products of these cDNAs, together with those of cDNAs previously cloned from rainbow trout (Oncorhynchus mykiss) and gilthead sea bream (Sparus aurata), were determined by heterologous expression in the yeast Saccharomyces cerevisiae. The carp and turbot desaturase cDNAs included open reading frames (ORFs) of 1335 and 1338 base pairs, respectively, specifying proteins of 444 and 445 amino acids. The protein sequences possessed all the characteristic features of microsomal fatty acid desaturases, including three histidine boxes, two transmembrane regions, and N-terminal cytochrome b(5) domains containing the haem-binding motif, HPGG. Functional expression showed all four fish cDNAs encode basically unifunctional Delta6 fatty acid desaturase enzymes responsible for the first and rate-limiting step in the biosynthesis of HUFA from 18:3n-3 and 18:2n-6. All the fish desaturases were more active towards the n-3 substrate with 59.5%, 31.5%, 23.1% and 7.0% of 18:3n-3 being converted to 18:4n-3 in the case of turbot, trout, sea bream and carp, respectively. The enzymes also showed very low, probably physiologically insignificant, levels of Delta5 desaturase activity, but none of the products showed Delta4 desaturase activity. The cloning and characterization of desaturases from these fish is an important advance, as they are species in which there is a relative wealth of data on the nutritional regulation of fatty acid desaturation and HUFA synthesis, and between which substantive differences occur.
鱼类是n-3高度不饱和脂肪酸（HUFA）、二十碳五烯酸（EPA）和二十二碳六烯酸（DHA）最重要的膳食来源，这些脂肪酸在人体营养中具有特别重要的作用，反映了它们在关键生理过程中的作用。本文所述研究的目的是克隆、功能表征和比较淡水和海水硬骨鱼类中参与EPA和DHA生成的脂肪酸去饱和酶基因。从鲤鱼（Cyprinus carpio）和大菱鲆（Psetta maximus）中分离并克隆了推定的脂肪酸去饱和酶cDNA。通过在酿酒酵母中进行异源表达，测定了这些cDNA产物以及先前从虹鳟（Oncorhynchus mykiss）和金头鲷（Sparus aurata）中克隆的cDNA产物的酶活性。鲤鱼和大菱鲆的去饱和酶cDNA分别包含1335和1338个碱基对的开放阅读框（ORF），分别编码444和445个氨基酸的蛋白质。蛋白质序列具有微粒体脂肪酸去饱和酶的所有特征，包括三个组氨酸框、两个跨膜区域以及含有血红素结合基序HPGG的N端细胞色素b(5)结构域。功能表达表明，所有四种鱼类cDNA编码的基本上是单功能的Δ6脂肪酸去饱和酶，负责从18:3n-3和18:2n-6生物合成HUFA的第一步和限速步骤。所有鱼类去饱和酶对n-3底物的活性更高，在大菱鲆、虹鳟、金头鲷和鲤鱼中，分别有59.5%、31.5%、23.1%和7.0%的18:3n-3转化为18:4n-3。这些酶还表现出非常低的、可能在生理上无显著意义的Δ5去饱和酶活性水平，但没有任何产物表现出Δ4去饱和酶活性。从这些鱼类中克隆和表征去饱和酶是一项重要进展，因为它们是在脂肪酸去饱和和HUFA合成的营养调控方面有相对丰富数据且存在实质性差异的物种。
