植物脂氧合酶研究进展

脂氧合酶（简称LOX）是广泛分布的含有非血红素离子的双加氧酶,它是植物十八碳酸代谢途径的关键酶．该途径也称LOX途径。因为该酶作用的产物在植物的生长发育过程中以及在植物对环境胁迫反应中起着重要的作用,因此一直是人们研究的热点。目前对于植物脂氧舍酶的研究主要集中在脂氧合酶基因的表达调控、LOX途径的生化研究以及代谢产物的生理功能这几个方面。     

乳突果苗的组织培养及脂氧合酶的诱导

脂氧合酶(LOX)是植物十八碳酸途径中一个很重要的酶,该酶作用的产物在植物生长发育过程以及在植物对环境胁迫反应中起着重要作用。首次建立了萝藦科植物乳突果苗的培养体系,并用仿真菌环境(几丁质)对乳突果组培苗进行刺激诱导,通过LC-ESI-MS检测脂氧合酶反应的产物。粗酶活性鉴定结果显示,经150mg/L几丁质诱导9h的乳突果组培苗产生了9-LOX,该酶可催化亚油酸生成9,10,11-三羟基-12-十八碳烯酸。推测在乳突果组培苗中,几丁质诱导的十八碳酸代谢途径沿着9-LOX方向进行。     

钙以及钙和钙调素拮抗剂对小麦幼苗LOX活性的影响（简报）

施用外源钙、EGTA和TFP,研究外源钙及钙和钙调素拮抗剂对小麦幼苗脂氧合酶活性的影响,结果表明,外源钙抑制脂氧合酶活性;正常生长的植物组织中的钙水平已使脂氧合酶处于一定程度的抑制状态;脂氧合酶并不是一种Ca2＋-CaM依赖的酶。     

贝莱斯芽孢杆菌生防次级代谢产物研究进展

贝莱斯芽孢杆菌（Bacillus velezensis）是生防芽孢杆菌中的重要代表,作为微生物农药的核心菌种,已被广泛应用于作物病害生物防治。贝莱斯芽孢杆菌具有植物内生性,其生防作用机制主要包括产生次级代谢产物对抗植物病原物;改善宿主植物根际微生物群落,促进宿主营养和生长;激发宿主植物产生防御反应,诱导植物获得系统抗性。其中,产生次级代谢产物是其最重要的生防作用机制。贝莱斯芽孢杆菌含有多个编码生物合成次级代谢产物的基因簇,其中包括编码聚酮化合物合酶（PKS）和非核糖体肽合成酶（NRPS）的基因簇,同时存在核糖体途径合成次级代谢产物基因簇。通过非核糖体途径可产生脂肽类化合物、聚酮类化合物、二肽和铁载体;通过核糖体途径产生小菌素、细菌素、羊毛硫抗生素。这些具有生物活性的次级代谢产物成为了天然新药和候选抗生素的储存库,对于解析生防菌作用机制具有重要意义。本文综述了贝莱斯芽孢杆菌的命名与更迭,产生次级代谢产物的类型、合成与调控基因以及靶标病原菌,以期为生防菌株的改良和生物农药的研发提供参考。     

植物鞘脂的结构、代谢途径及其功能

众所周知, 鞘脂是生物膜结构的重要组成成分, 随着鞘脂在动物和酵母中的深入研究发现, 鞘脂及其代谢产物是一类很重要的活性分子, 它们参与调节细胞的生长、分化、衰老和细胞程序性死亡等许多重要的信号转导过程。鞘脂在植物中的研究最近几年才开始, 植物鞘脂的功能还不十分清楚。最近的研究发现, 鞘脂及其代谢产物在植物中也起着很重要的信号分子作用。该文详细总结了鞘脂在植物中的结构、代谢途径和主要生物学功能, 并结合实验室的工作对植物鞘脂的功能研究进行了展望。     

植物鞘脂的结构、代谢途径及其功能

众所周知,鞘脂是生物膜结构的重要组成成分,随着鞘脂在动物和酵母中的深入研究发现,鞘脂及其代谢产物是一类很重要的活性分子,它们参与调节细胞的生长、分化、衰老和细胞程序性死亡等许多重要的信号转导过程。鞘脂在植物中的研究最近几年才开始,植物鞘脂的功能还不十分清楚。最近的研究发现,鞘脂及其代谢产物在植物中也起着很重要的信号分子作用。该文详细总结了鞘脂在植物中的结构、代谢途径和主要生物学功能,并结合实验室的工作对植物鞘脂的功能研究进行了展望。     

蜡酯合成途径及关键酶的研究进展

蜡酯对于生物的生命活动具有重要意义,研究表明植物和动物的蜡酯合成存在保守途径。即脂酰辅酶A（fatty acyl-CoA）在脂酰辅酶A还原酶（fatty acyl-CoAreductase,FAR）的作用下还原成脂肪醇,脂肪醇和脂酰辅酶A在蜡酯合酶（wax synthase,WS）的作用下生成酯,FAR和WS是该途径的关键酶,这两个酶的结构和功能在不同物种之间表现出很大差异,目前对于这两个酶缺乏系统的归纳分析。该文综述了蜡酯合成途径及FAR和WS的序列特征、生化特性及参与的生理功能,分析了这两种酶相关研究存在的问题,旨在为昆虫的蜡酯合成研究提供参考。     

虫害诱导的植物挥发物代谢调控机制研究进展

长期受自然界的非生物/生物侵害,植物逐步形成了复杂的防御机制,为防御植食性昆虫的为害,植物释放虫害诱导产生的挥发性化合物(herbivore-induced plant volatiles,HIPVs)。HIPVs是植物-植食性昆虫-天敌三级营养关系之间协同进化的结果。HIPVs的化学组分因植物、植食性昆虫种类的不同而有差异。生态系统中,HIPVs可在植物与节肢动物、植物与微生物、虫害植物与邻近的健康植物、或同一植株的受害和未受害部位间起作用,介导防御性反应。HIPVs作为寄主定位信号,在吸引捕食性、寄生性天敌过程中起着重要作用。HIPVs还可以作为植物间信息交流的工具,启动植株的防御反应而增强抗虫性。不论从生态学还是经济学角度来看,HIPVs对于农林生态系中害虫综合治理策略的完善具有重要意义。前期的研究在虫害诱导植物防御的化学生态学方面奠定了良好基础,目前更多的研究转向阐述虫害诱导植物抗性的分子机制。为了深入了解HIPVs的代谢调控机制,主要从以下几个方面进行了综述。因为植食性昆虫取食造成的植物损伤是与昆虫口腔分泌物共同作用的结果,所以首先阐述口腔分泌物在防御反应中的作用。挥发物诱导素volicitin和β-葡萄糖苷酶作为口腔分泌物的组分,是产生HIPVs的激发子,通过调节伤信号诱发HIPVs的释放。接着阐述了信号转导途径对HIPVs释放的调节作用,并讨论了不同信号途径之间的交互作用。就HIPVs的代谢过程而言,其过程受信号转导途径(包括茉莉酸、水杨酸、乙烯、过氧化氢信号途径)的调控,其中茉莉酸信号途径是诱发HIPVs释放的重要途径。基于前人的研究,综述了HIPVs的主要代谢过程及其过程中关键酶类的调控作用。文中的HIPVs主要包括萜烯类化合物、绿叶挥发物和莽草酸途径产生的芳香族化合物,如水杨酸甲酯和吲哚等。作为化学信号分子,这些化合物中的一部分还能激活邻近植物防御基因的表达。萜烯合酶是各种萜烯类化合物合成的关键酶类,脂氧合酶、过氧化氢裂解酶也是绿叶挥发物代谢途径中的研究热点,而苯丙氨酸裂解酶和水杨酸羧基甲基转移酶分别是合成水杨酸及其衍生物水杨酸甲酯的关键酶类。这些酶类的基因在转录水平上调控着HIPVs代谢途径。最后展望了HIPVs的研究前景。     

植物维生素E合成相关酶基因的克隆及其在体内功能研究进展

维生素E（VE）的天然产物有8种类型,分别为α、β、γ、δ-生育酚（tocopherol）和α、β、γ、δ-生育三烯酚（tocotrienol）,对植物、动物和人类都具有十分重要的生理作用。医学证明,维生素E不仅与生殖系统,而且与中枢神经系统、消化系统、心血管系统和肌肉系统的正常代谢都有密切关系;它也是治疗冠心病、动脉粥样硬化、贫血、脑软化、肝病和癌症等的辅助药物。而绿色植物则是人类和动物VE的基本来源。近年来随着植物基因组学的发展和营养基因组学概念的提出,通过快速分离植物营养代谢（VE）相关酶的基因,最终解析和调控植物微量营养素代谢途径,利用代谢工程的方法大大提高植物营养价值之策略正在逐步完善。本文对有关植物中VE生物合成途径和相关酶基因克隆研究现状,以及VE在植物体内的作用和功能研究进展进行了综述,以期为VE作用机理的探寻和功能开发提供新思路。     

5-脂氧合酶抑制剂的研究进展

脂氧合酶是一种含铁的氧化还原酶，存在于动植物的组织和细胞中。脂氧合酶主要分为1、3、5、8、12、15-脂氧合酶6种亚型。5-脂氧合酶(5-lipoxygenase,5-LOX)是LOX同工酶中研究较为深入的非血红素铁蛋白氧化代谢酶，是许多高活性氧化脂质生物合成中的关键酶，参与多种炎症性疾病，如炎症、哮喘、脑缺血、部分恶性肿瘤及阿尔茨海默症等疾病的发生和发展过程。因此，开发有效的5-LOX抑制剂对治疗上述疾病至关重要。本文介绍了5-LOX抑制剂的研究成果，并对其分类、来源、治疗药效和优缺点进行了讨论，为深入研发创新型5-LOX抑制剂提供理论依据。     

Identification of putative residues involved in the accessibility of the substrate-binding site of lipoxygenase by site-directed mutagenesis studies

Lipoxygenases (LOXs) are a class of widespread dioxygenases catalyzing the hydroperoxidation of polyunsaturated fatty acids (PUFA). Recently, we isolated a cDNA encoding a LOX, named olive LOX1, from olive fruit of which the deduced amino acid sequence shows more than 50% identity with plant LOXs. In the present study, a model of olive LOX1 based on the crystal structure of soybean LOX-1 as template has been generated and two bulky amino acid residues highly conserved in LOXs (Phe277) and in plant LOXs (Tyr280), located at the putative entrance of catalytic site were identified. These residues may perturb accessibility of the substrate-binding site and therefore were substituted by less space-filling residues. Kinetic studies using linoleic and linolenic acids as substrates were carried out on wild type and mutants. The results show that the removal of steric bulk at the entrance of the catalytic site induces an increase of substrate affinity and of catalytic efficiency, and demonstrate that penetration of substrates into active site of olive LOX1 requires the movement of the side chains of the Phe277 and Tyr280 residues. This study suggests the involvement of these residues in the accessibility of the substrate-binding site in the lipoxygenase family.     

Analysis of the subcellular localisation of lipoxygenase in legume and actinorhizal nodules

Plant lipoxygenases (LOXs; EC 1.13.11.12) catalyse the oxygenation of polyunsaturated fatty acids, linoleic (18:2) and α-linolenic acid (18:3(n-3)) and are involved in processes such as stress responses and development. Depending on the regio-specificity of a LOX, the incorporation of molecular oxygen leads to formation of 9- or 13-fatty acid hydroperoxides, which are used by LOX itself as well as by members of at least six different enzyme families to form a series of biologically active molecules, collectively called oxylipins. The best characterised oxylipins are the jasmonates: jasmonic acid (JA) and its isoleucine conjugate that are signalling compounds in vegetative and propagative plant development. In several types of nitrogen-fixing root nodules, LOX expression and/or activity is induced during nodule development. Allene oxide cyclase (AOC), a committed enzyme of the JA biosynthetic pathway, has been shown to localise to plastids of nodules of one legume and two actinorhizal plants, Medicago truncatula, Datisca glomerata and Casuarina glauca, respectively. Using an antibody that recognises several types of LOX interspecifically, LOX protein levels were compared in roots and nodules of these plants, showing no significant differences and no obvious nodule-specific isoforms. A comparison of the cell-specific localisation of LOXs and AOC led to the conclusion that (i) only cytosolic LOXs were detected although it is generally assumed that the (13S)-hydroperoxy α-linolenic acid for JA biosynthesis is produced in the plastids, and (ii) in cells of the nodule vascular tissue that contain AOC, no LOX protein could be detected.     

植物脂肪氧化酶的研究进展

植物脂肪氧化酶(LOX)是一个多基因家族, 是由单一的多肽链组成的含有非血红素铁、不含硫的过氧化物酶。LOX催化具有顺, 顺-1, 4-戊二烯结构的多元不饱和脂肪酸的双加氧反应。植物中, 不同脂肪氧化酶的最适pH、pI、底物和产物特异性、时空表达特性、亚细胞定位等存在差异。LOX参与的生理过程涉及损伤、病原攻击、种子萌芽、果实熟化、植物衰老、脱落酸和茉莉酸合成, LOX也在正常的植物生长和生殖生长过程中作为营养储藏蛋白, 参与脂类迁移、响应营养胁迫、调节“源”与“库”的分配。对LOX家族的深入理解,将有助于LOX在作物新品种的选育、新型植保素的开发、食品加工等方面得到广泛的应用。     

Metabolic profiling of oxylipins upon sorbitol treatment in barley leaves

In barley leaves 13-lipoxygenases (LOXs) are induced by salicylate and jasmonate. Here, we analyse by metabolic profiling the accumulation of oxylipins upon sorbitol treatment. Although 13-LOX-derived products are formed and specifically directed into the reductase branch of the LOX pathway, accumulation is much later than in the cases of salicylate and jasmonate treatment. In addition, under these conditions only the accumulation of jasmonates as additional products of the LOX pathway has been found.     

Biochemical and molecular characterization of hazelnut (Corylus avellana) seed lipoxygenases.

Plant lipoxygenases (LOXs) are a class of dioxygenases which display diverse functions in several physiological processes such as growth, development and response to biotic and abiotic stresses. Even though LOXs have been characterized from several plant species, the physiological role of seed LOXs is still unclear. With the aim to better clarify the occurrence of LOXs and their influence on hazelnut seed quality, we carried out the biochemical and molecular characterization of the main LOX isoforms expressed during seed development. A genomic clone containing a complete LOX gene was isolated and fully characterized. The 9887 bp sequence reported contains an open reading frame of 5334 bp encoding a putative polypeptide of 99 kDa. Semiquantitative RT-PCR carried out from RNAs extracted from seeds at different maturation stages showed that LOXs are mainly expressed at early developmental stages. These results were confirmed by LOX activity assays. Biochemical characterization of the reaction products of the hazelnut LOX indicated that it is a 9-LOX. Two cDNAs were isolated by RT-PCR carried out on total RNA from immature hazelnut seeds. Sequence analysis indicated that the two cDNAs are highly homologous (91.9% degree of identity) and one of these corresponded exactly to the genomic clone. The deduced amino acid sequences of the hazelnut LOXs showed that they are closely related to a previously reported almond LOX (79.5% identity) and, to a lesser extent, to some LOXs involved in plant responses to pathogens (cotton and tobacco LOXs, 75.5 and 74.6% identity, respectively). The physiological role of hazelnut LOXs and their role in influencing seed quality are also discussed.     

Cunxi Wang · Kevan P.C. Croft · David F. Hildebrand

 Auxin [α-naphthaleneacetic acid (NAA) or indole-3-acetic acid] can induce the expression of lipoxygenases (LOXs) in cultured immature zygotic embryo cotyledons of soybean [Glycine max. (L.) Merr]. These auxin-induced LOXs are different from those normally expressed in seeds but have the same isoelectric points (pI) as those found in seedlings. The pIs of the two seedling LOXs were determined to be 5.09 and 5.23. One of the auxin-induced LOXs has the same pI (5.09) and molecular mass (94 kDa) as seedling LOX4. The partial amino acid sequences from the purified NAA-induced pI-5.09 LOX are identical to those of LOX4. RNA protection assays showed that NAA induces the expression of LOX4 and LOX5 mRNAs in cultured embryo cotyledons where they are not normally expressed. Soybean genotypes with a polymorphic variant of LOX4 in hypocotyls showed the same variation as NAA-induced LOXs in the embryo cotyledons. These results demonstrate that the NAA-induced pI-5.09 LOX is seedling LOX4 and also suggest that auxin might be directly or indirectly involved in seedling LOX expression during seed germination. Received: 10 January 2000 / Revision received: 16 June 2000 / Accepted: 29 June 2000     

Industrial potential of lipoxygenases

Lipoxygenases (LOXs) are iron- or manganese-containing oxidative enzymes found in plants, animals, bacteria and fungi. LOXs catalyze the oxidation of polyunsaturated fatty acids to the corresponding highly reactive hydroperoxides. Production of hydroperoxides by LOX can be exploited in different applications such as in bleaching of colored components, modification of lipids originating from different raw materials, production of lipid derived chemicals and production of aroma compounds. Most application research has been carried out using soybean LOX, but currently the use of microbial LOXs has also been reported. Development of LOX composition with high activity by heterologous expression in suitable production hosts would enable full exploitation of the potential of LOX derived reactions in different applications. Here, we review the biological role of LOXs, their heterologous production, as well as potential use in different applications. LOXs may fulfill an important role in the design of processes that are far more environmental friendly than currently used chemical reactions. Difficulties in screening for the optimal enzymes and producing LOX enzymes in sufficient amounts prevent large-scale application so far. With this review, we summarize current knowledge of LOX enzymes and the way in which they can be produced and applied.