CAJ | 학술논문

脂肪酸延长酶5(ELOVL5)是高不饱和脂肪酸(HUFA)合成的关键酶之一。为了研究鲤HUFA合成的能力和机制,本研究采用RT-PCR和RACE技术获得镜鲤ELOVL5全长cDNA序列。ELOVL5基因cDNA全长1121 bp,开放阅读框为876 bp,编码291个氨基酸。序列分析显示,镜鲤ELOVL5氨基酸序列包含1个组氨酸簇(HXXHH),1个典型的内质网驻留信号,多个跨膜区域和多个保守区域(KXXEXXDT、QXXFLHXYHH、NXXXHXXMYXYY、TXXQXXQ),具有典型的脂肪酸延长酶的结构特征。氨基酸同源性分析结果显示,镜鲤 ELOVL5基因与其他鱼类同源性为79.0%~93.1%,与人同源性为69.0%。通过实时荧光定量PCR(RT-qPCR)检测该基因在镜鲤不同组织中的表达量,发现脂肪酸延长酶基因在镜鲤肝中表达量最高,其次为脑,在背部肌肉中表达量最低。镜鲤ELOVL5基因的获得为进一步研究镜鲤HUFA的合成途径及调控机理奠定了基础。
지방산연장매5(ELOVL5)시고불포화지방산(HUFA)합성적관건매지일。위료연구리HUFA합성적능력화궤제,본연구채용RT-PCR화RACE기술획득경리ELOVL5전장cDNA서렬。ELOVL5기인cDNA전장1121 bp,개방열독광위876 bp,편마291개안기산。서렬분석현시,경리ELOVL5안기산서렬포함1개조안산족(HXXHH),1개전형적내질망주류신호,다개과막구역화다개보수구역(KXXEXXDT、QXXFLHXYHH、NXXXHXXMYXYY、TXXQXXQ),구유전형적지방산연장매적결구특정。안기산동원성분석결과현시,경리 ELOVL5기인여기타어류동원성위79.0%~93.1%,여인동원성위69.0%。통과실시형광정량PCR(RT-qPCR)검측해기인재경리불동조직중적표체량,발현지방산연장매기인재경리간중표체량최고,기차위뇌,재배부기육중표체량최저。경리ELOVL5기인적획득위진일보연구경리HUFA적합성도경급조공궤리전정료기출。
Highly unsaturated fatty acids (HUFAs), such as eicosapentaenoic acid and docosahexaenoic acid, are essen-tial for normal growth and development and play an important role in the prevention and treatment of coronary artery disease, hypertension, arthritis and cancer. Enzymes that lengthen the carbon chain of polyunsaturated fatty acids are vital to biosynthesis of the highly unsaturated fatty acids, arachidonic acid, eicosapentaenoic acid and docosahexaenoic acid from linoleic and a-linolenic acids. Fatty acid elongase 5 (ELOVL5) is a member of the ELOVL family and is mainly responsible for 18 carbon-carbon chain extension. To study the regulatory role of ELOVL5 in biosynthesis of HUFA, the full-length ELOVL5 cDNA from mirror carp was cloned using reverse transcriptase PCR and rapid ampli-fication of cDNA ends. The ELOVL5 cDNA was 1 121 bp, with an open reading frame of 876 bp, encoding a protein of 291 amino acids containing a single histidine box, a canonical ER retention signal and several transmembrane regions, as seen in other fatty acid elongases. The protein shares 79.0%–93.1% sequence identity with other fish, and 69.0%identity with Homo sapiens. The phylogenetic tree showed that it clustered closely with herbivorous and omnivorous freshwater fish. ELOVL5 gene expression in various mirror carp tissues was determined using real-time quantitative PCR. Expression was highest in liver, followed by brain, and lowest in muscle. The fact that the ELOVL5 expression level was highest in brain and liver than in other tissues indicates that these are the main tissues for HUFA biosynthesis. We found the ELOVL5 expression level lowest in muscle and this may be related to its physiological function in this tissue. These results will assist understanding of the biosynthesis of HUFA in common carp, and enable methods to be developed for enhancing its production. Enhanced HUFA production will accelerate the development and application of formulated feeds, and assist the development of aquiculture.