植物纤维素合酶基因研究进展

纤维素合酶催化合成的β_1,4糖苷链构成植物细胞壁中含量最丰富的组份纤维素。植物体中存在着众多纤维素合酶,同时还具多种与之相关的纤维素合酶相似蛋白,它们组成了一个庞大的纤维素合酶超家族。纤维素合酶的催化机理尚不清楚,纤维素合酶相似蛋白的功能更有待于深入研究。本文综述了近年植物纤维素合酶及其相似蛋白编码基因的研究进展。     

高等植物纤维素合酶超家族

从1996年第一个植物纤维素合酶基因的鉴定,人们对植物体内纤维素合成的研究已经走过了10年的历程。10多年中,人们取得了很大的成果,也有很多问题有待解决。该文主要介绍拟南芥和毛果杨基因组中的纤维素合酶超家族。     

苎麻纤维素合酶BnCesA1高变区蛋白的原核表达与条件优化

高等植物纤维素合酶是含有多个功能结构域的糖苷转移酶,催化合成β-1 ,4葡萄糖苷链,即高等植物细胞壁的重要成分之一纤维素。在同一物种例如在拟南芥中比对纤维素合酶家族各成员的序列信息,发现其蛋白序列中存在两个高变异区域（HVR）,其中第一个HVR靠近NH2端（NHVR）,富含酸性氨基酸。本研究克隆了苎麻纤维素合酶基因（BnCesA1） 的NHVR编码序列,并以正确的阅读框亚克隆至含有6×His接头的pQE-N1原核表达载体,构建了pQE-N1-NHVR重组表达载体。在大肠杆菌BL21（DE3）中表达的重组蛋白His-tag-NHVR经Western-blotting验证后,正交优化得到该蛋白小量表达的最优条件组合为：所挑取的2号菌落在37 ℃下,IPTG的诱导浓度为0.1 mmol/L, 诱导表达4 hr。本研究结果为纯化His-tag-NHVR融合蛋白,制备其抗体及进一步研究苎麻纤维素合酶局部功能或其组织特异性作用打下基础。     

植物细胞壁中纤维素合成的研究进展

纤维素是植物细胞壁的主要成分,是植物细胞壁执行生理功能的基础,也是人类生产和生活中必不可少的一类物质。本文对纤维素合成、合成中所需要的酶以及纤维素沉积中微纤丝的作用等方面进行了综述和探讨, 并对纤维素合成的深入研究进行了展望。     

蔗糖合酶在植物生长发育中的作用研究

蔗糖合酶（SuSy）是植物蔗糖代谢的关键酶之一,在植物各组织中普遍存在。SuSy参与了植物体中许多代谢过程,包括淀粉及纤维素的合成,以及碳源的分配等。该酶还可影响植物的抗逆性、种子发育和生物固氮能力,因此,利用SUS基因改良作物品质具有良好的应用前景。对SuSy的性质、基因表达模式及其在植物生长发育中的作用进行综述。     

苎麻纤维素合酶基因BnCesA1全长cDNA的克隆与表达分析

苎麻是中国的传统纤维作物,能够生产最长的自然纤维。本研究旨在克隆苎麻纤维素合酶基因BnCesA1全长编码序列,对其表达模式进行分析。以已知的苎麻纤维素合酶基因序列(DQ077190)为基础,设计5’RACE引物,以湘苎三号为材料,得到了BnCesA1的5’端,拼接后得到了BnCesA1的全长序列,并从湘苎三号的cDNA中成功克隆到包括BnCesA1全部编码序列的cDNA序列。扩增得到的BnCesA1基因cDNA为3253bp,编码区3246bp,编码含1082个氨基酸的多肽。通过对这个基因进行核酸序列和蛋白结构域分析表明,BnCesA1和毛果杨、欧美山杨、巨桉、大叶相思等其他物种的纤维素合酶基因都有很高的同源性,根据得到的BnCesA1的5’端设计特异性表达检测引物,分析其在湘苎三号苎麻品种各组织中的表达情况,结果显示BnCesA1在所检测的各组织中均有表达,表达量为茎皮>叶>顶芽>根。本研究首次克隆到苎麻中编码全长蛋白的纤维素合酶基因,并且苎麻BnCesA1在茎皮中高表达提示该基因可能在苎麻韧皮纤维合成中有重要作用。     

拟南芥纤维素合酶基因时空表达模式与功能预测

纤维素是细胞壁的主要组成成分, 研究纤维素合成可以从源头上解决关于高效降解纤维素的问题。该研究通过综合拟南芥(Arabidopsis thaliana)纤维素合酶基因(AtCESA)家族的进化和芯片表达分析及根据拟南芥全生育期GUS染色结果分析纤维素合酶基因的时空表达模式, 发现拟南芥纤维素合酶基因AtCESA1, 3, 6以及AtCESA4, 7, 8分别参与细胞壁初生壁和次生壁的合成并存在明显的共表达现象。其中, AtCESA1, 3, 6在全生育期表达, AtCESA4, 7, 8主要在根、茎和叶脉等次生壁细胞中表达。AtCESA5和AtCESA6、AtCESA2和AtCESA9以及AtCESA1和AtCESA10等基因对均有基因重复作用。根据AtCESA家族基因表达模式和分子演化关系可以推测, AtCESA5对AtCESA6以及AtCESA9对AtCESA2可能分别存在功能冗余。此外, AtCESA9的表达具明显的组织特异性。上述研究结果为深入认识拟南芥纤维素合酶基因的功能奠定了基础。     

毛白杨蔗糖合酶基因PtSUS1的克隆及其表达模式分析

蔗糖合酶（sucrose synthase）与植物库强调节、次生壁的形成和纤维素合成等有着密切的联系,其中在纤维素合成过程中的作用尤为显著。本研究根据我们已获得的毛白杨PtSUS1基因片段设计引物,采用RACE技术,获得了毛白杨PtSUS1的基因序列,测序结果显示该基因序列全长为2 669 bp,包括一个完整的阅读框,编码805个氨基酸。通过Blast检索分析表明,PtSUS1与拟南芥、巨桉、陆地棉、温州蜜柑、毛果杨SUS1的核酸和氨基酸序列的同源性分别达到76%~97%和82%~97%。运用生物信息软件对PtSUS1编码的蛋白进行了二级结构预测和功能位点分析,结果显示该蛋白氨基酸序列包括两个功能域,存在可能的磷酸化位点38个,无跨膜结构域存在。系统进化分析表明PtSUS1与PtSUS2关系最为接近。RT-PCR分析结果显示,PtSUS1在被检测的毛白杨根、茎、叶及雌雄花芽组织和器官中均有表达,呈现组成型表达模式。该研究为进一步深入探索毛白杨蔗糖合酶基因PtSUS1的功能奠定了基础。     

萌发后食松幼苗淀粉粒结合与可溶性淀粉合酶的研究

裸子植物食松 (PinusedulisEngelm .)幼苗在萌发后具有淀粉积累的独特特征 ,而淀粉合酶在裸子植物中尚未有研究。这项研究对在室温下暗中萌发的食松幼苗的淀粉合酶进行了提取和研究 ,结果显示 :5 8kD和 91kD的淀粉粒结合蛋白均为豌豆淀粉合酶Ⅱ抗体所识别。5 8kD淀粉粒结合蛋白被纯化出来 ,其N端序列与其他几种被子植物淀粉粒结合的淀粉合酶Ⅰ的N端序列有很高的相似性 ,并与豌豆、大麦、马铃薯淀粉粒结合淀粉合酶Ⅰ有免疫相似性 ,被鉴定为淀粉粒结合的淀粉合酶Ⅰ。从DEAE_Sepharose柱上洗脱出来的可溶性淀粉合酶表现出两个淀粉合酶活性峰 ,表明至少有两类可溶性淀粉合酶同工酶。对可溶性淀粉合酶的前体亲和特性进行了研究。以兔肝糖原为前体时 ,可溶性淀粉合酶的亲和性最高。裸子植物淀粉合酶的酶学特性表现与已研究的被子植物相似。     

萌发后食松幼苗淀粉粒结合与可溶性淀粉合酶的研究

裸子植物食松(Pinus edulis Engelm.)幼苗在萌发后具有淀粉积累的独特特征,而淀粉合酶在裸子植物中尚未有研究.这项研究对在室温下暗中萌发的食松幼苗的淀粉合酶进行了提取和研究,结果显示: 58 kD和91 kD的淀粉粒结合蛋白均为豌豆淀粉合酶Ⅱ抗体所识别. 58 kD淀粉粒结合蛋白被纯化出来, 其N端序列与其他几种被子植物淀粉粒结合的淀粉合酶Ⅰ的N端序列有很高的相似性,并与豌豆、大麦、马铃薯淀粉粒结合淀粉合酶Ⅰ有免疫相似性,被鉴定为淀粉粒结合的淀粉合酶Ⅰ.从DEAE-Sepharose 柱上洗脱出来的可溶性淀粉合酶表现出两个淀粉合酶活性峰,表明至少有两类可溶性淀粉合酶同工酶.对可溶性淀粉合酶的前体亲和特性进行了研究.以兔肝糖原为前体时, 可溶性淀粉合酶的亲和性最高.裸子植物淀粉合酶的酶学特性表现与已研究的被子植物相似.     

Green composites from sustainable cellulose nanofibrils: A review

Green composites are materials having ecofriendly attributes that are technically and economically feasible while minimizing the generation of pollution. In this context it refers to the combination of fully degradable fibers mostly cellulosic materials and natural resins to develop green composite materials. In the past decade, overdependence on petroleum products (synthetic polymers, resins, etc.) has consistently increased and on account of this, the researchers are now focusing more on green materials specially cellulosics. Cellulosic fibers in micro and nano scale are attractive to replace man-made fibers as reinforcement to make environmentally friendly green products. In this study, we will discuss the processing, extraction, properties, chronological events and applications of cellulose and cellulosic-based nanocomposite materials. Cellulosic nanocomposites are currently considered one of the most promising areas of scientific and technological development in the field of plant products. The aim of this review is to demonstrate the current state of development in the field of cellulose nanofibril based green composites research and application through examples.     

Crisis in Biology An Examination of the Deficiencies in the Current Pattern of Expansion in Biological Education

A method of analysing the content of biology curricula, courses, and texts on the basis of the relative emphasis laid on different levels of biological organization, is discussed. It is suggested that this method may have led to unwarranted contrasts between ‘traditional’ and ‘modern’ curricula. A simple modification to this method is proposed.     

微生物降解纤维素的新机制

自然界中能够降解纤维素的微生物分布广泛,纤维素降解的方式也呈现很高的多样性。Cytophaga hutchinsonii属于拟杆菌门,具有很强的结晶纤维素降解能力,但是其降解机制既异于已经发现的游离纤维素酶降解体系,也不同于纤维小体的降解模式,推测其存在第3种细胞结合型纤维素降解模式。本文简要阐述以C. hutchinsonii为代表的一种新的纤维素降解策略。     

Recent progress in cellulose biosynthesis

Cellulose comprises the major polymer of the plant cell wall. It consists of a set of parallel chains composed of glucans and these chains are highly oriented to form a structure known as a microfibril. The orientation of the microfibrils controls the extension of the direction of the plant cell. Extensive studies on the cellulose biosynthesis have been carried out for over three decades, and recently (1996) genes for cellulose biosynthesis in plants (CesA) were isolated. In the year 2002, a specific primer for cellulose biosynthesis reaction has been discovered and cellulose synthetic activity has been also confirmed by recombinant protein derived from the plant CesA gene. Furthermore, other proteins involved in cellulose biosynthesis besides CesA proteins were also proposed at the same time. One of these proteins, Korrigan cellulase, was suggested to act by removing sitosterol from the primer for biosynthesis reaction of cellulose. A membrane-bound sucrose synthase was also suggested to provide UDP-glucose as a substrate for cellulose biosynthesis. On the basis of these results, a new pathway for cellulose biosynthesis was proposed. Now, the research field of cellulose biosynthesis is facing a major turning point. Electronic Publication     

纤维素降解菌的筛选与复合菌系的初步构建

目的针对大庆地区土地盐碱化较严重,盐碱面积较大等现状,从大庆特征性盐碱地区土壤样品中分离出能够降解纤维素的菌株,为降解植物废料以及缓解土壤盐碱化改善土壤环境提供功能菌株。方法通过刚果红染色法初步筛得到具有纤维素降解能力菌株,进一步用比色法测定其纤维素降解率,同时测定菌株耐盐、耐碱、产酸性能,选择性能优良的3株菌作为纤维素降解菌复合菌系构建菌株,通过耐盐性、耐碱性和纤维素降解率实验测定复合菌系菌株最佳组合,进一步通过上述实验确定复合菌系菌株最佳混合比例。结果得到由DX-5和DX-9按照2∶3进行组合复合菌系纤维素降解率最高,达到87.96%,且具有较高的耐盐以及耐碱能力。结论通过实验得到纤维素降解菌的复合菌系,具有较高纤维素降解率以及改善土壤盐碱性能力。     

纤维素酶促进剂在饲料中的应用初探

采用3.5-二硝基水杨酸(DNS)比色法和重量法,分别研究纤维酶促进剂在鸡饲料中使酶活力提高的作用,取得良好效果。对不同来源的纤维素酶的扩大实验,进一步证明了促进剂的有效性和普遍性,确定了纤维素酶促进剂应用于饲料的最佳配比,证明了不同促进剂之间存在明显的协同促进作用。动物体外模拟实验使促进剂更加实用化,结果表明:纤维素酶促进剂可提高纤维素酶的酶解率,降低饲料成本,提高饲料利用率。     

Synthesis of cellulose dehydroabietate in ionic liquid [bmim]Br

A new type of cellulose derivative, cellulose dehydroabietate (CDA), was synthesized by the O-acylation reaction of cellulose with dehydroabietic acid chloride (DHAC) using ionic liquid 1-butyl-3-methylimidazolium bromide ([bmim]Br) as a solvent and 4-dimethyl-aminopyridine (DMAP) as a catalyst. The resulting CDA was characterized by means of FT-IR, X-ray diffraction (XRD), scanning electron microscopy (SEM), thermal gravimetric analysis (TGA), and elemental analysis. Also, some properties of CDA were determined. These results showed that CDA has better solubility, water-repellency, and resistance to acids and bases than raw cellulose, and these properties increase with the DS of CDA.     

Comb-shaped graft copolymers with cellulose side-chains prepared via click chemistry

Comb-shaped copolymers with cellobiose acetate or cellulose triacetate (CTA) side-chains, PPMA-g-(CTA2-C15) and PPMA-g-(CTA13-C15), were prepared by grafting N-(15-azidopentadecanoyl)-2,3,6-tri-O-acetyl-4-O-(2,3,4,6-tetra-O-acetyl-β-d-glucopyranosyl)-β-d-glucopyranosylamine (CTA2-C15-N₃) and N-(15-azidopentadecanoyl)-tri-O-acetyl-β-cellulosylamine (CTA13-C15-N₃, number average degree of polymerization (DP_n) = 13) onto poly(2-propyn-1-yl methacrylate) (PPMA, weight average degree of polymerization (DP_w, X + Y = 5.59 × 10²)) via “click chemistry”. The copolymers were characterized by ¹H, ¹³C and two-dimensional NMR and size exclusion chromatography-multi-angle laser light scattering (SEC-MALS) measurements. The numbers of CTA side-chains (X) of PPMA-g-(CTA2-C15) and PPMA-g-(CTA13-C15) were calculated as 4.03 × 10² and 2.45 × 10², respectively. Copolymers with cellulosic side-chains, PPMA-g-(CELL2-C15) and PPMA-g-(CELL13-C15), were successfully obtained after deacetylation of PPMA-g-(CTA2-C15) and PPMA-g-(CTA13-C15), respectively. X-ray diffraction measurements revealed that PPMA-g-(CELL13-C15) showed crystalline pattern of cellulose II, which is believed to have anti-parallel orientation.     

On the polarity of cellulose in the cell wall of Valonia

The orientation of the triclinic phase of cellulose in the cell wall of Valonia ventricosa J. Agardh was investigated by X-ray- and electron-diffraction analysis. In addition to the well-documented uniplanar-axial organization of the cell wall which requires that the a ^* axis should be always perpendicular to the wall surface, the direction of this axis was also found to be pointing outward from the plasma membrane side of the wall. This unidirectionality was persistent throughout the various layers that constitute the cell wall and also for the three microfibrillar orientations that occur in Valonia cell walls. The unidirectionality of the a ^* axis indicates, in particular, that the Valonia cellulose microfibrils are not twisted along their axis. These observations are consistent with a cellulose biosynthetic scheme where a close association exists between terminal-complex orientations and those of the cellulose microfibrils. In this context, the unidirectionality of the a ^* axis of cellulose seems to be related to the restricted mobility of the terminal complexes which are able to slide in the plasma membrane but not to rotate along their long axis.Abbreviations TC terminal complex This work was initiated during a visit of J.F.R at Grenoble in the framework of a France-Québec exchange program. J.S. was recipient of a CNRS fellowship. The diagram in Fig. 8 was kindly drawn for us by Miss Yukie Saito from the Department of Forest Products, the University of Tokyo.