Receptivity To Free-Stream Disturbance Waves For Hypersonic Flow Over A Blunt Cone

A high-order shock-fitting finite difference scheme is studied and used to do direction numerical simulation (DNS) of hypersonic unsteady flow over a blunt cone with fast acoustic waves in the free stream, and the receptivity problem in the blunt cone hypersonic boundary layers is studied. The results show that the acoustic waves are the strongest disturbance in the blunt cone hypersonic boundary layers. The wave modes of disturbance in the blunt cone boundary layers are first, second, and third modes which are generated and propagated downstream along the wall. The results also show that as the frequency decreases, the amplitudes of wave modes of disturbance increase, but there is a critical value. When frequency is over the critial value, the amplitudes decrease. Because of the discontinuity of curvature along the blunt cone body, the maximum amplitudes as a function of frequencies are not monotone.

Effects Of The Length Of A Cylindrical Solid Shield On The Entrainment Of Ambient Air Into Turbulent And Laminar Impinging Argon Plasma Jets

When materials processing is conducted in air surroundings by use of an impinging plasma jet, the ambient air will be entrained into the materials processing region, resulting in unfavorable oxidation of the feedstock metal particles injected into the plasma jet and of metallic substrate material. Using a cylindrical solid shield may avoid the air entrainment if the shield length is suitably selected and this approach has the merit that expensive vacuum chamber and its pumping system are not needed. Modeling study is thus conducted to reveal how the length of the cylindrical solid shield affects the ambient air entrainment when materials processing (spraying, remelting, hardening, etc.) is conducted by use of a turbulent or laminar argon plasma jet impinging normally upon a flat substrate in atmospheric air. It is shown that the mass flow rate of the ambient air entrained into the impinging plasma jet cannot be appreciably reduced unless the cylindrical shield is long enough. In order to completely avoid the air entrainment, the gap between the downstream-end section of the cylindrical solid shield and the substrate surface must be carefully selected, and the suitable size of the gap for the turbulent plasma jet is appreciably larger than that for the laminar one. The overheating of the solid shield or the substrate could become a problem for the turbulent case, and thus additional cooling measure may be needed when the entrainment of ambient air into the turbulent impinging plasma jet is to be completely avoided.

Control Of Vortex Shedding From A Square Cylinder

Small circular, square, and thin-strip cross-sectional elements are used to suppress vortex shedding from a square cylinder at Reynolds numbers in the range of 1.12 x 10(4)-1.02 x 10(5). The axes of the element and cylinder are parallel. The element's size, position, and angle of attack are varied. Measurements of the fluctuating surface pressures and wake velocities, together with smoke flow visualization, show that vortex shedding from both sides of the cylinder is suppressed and the mean drag and fluctuating lift on the cylinder is reduced if the element is installed in an effective zone downstream of the cylinder. The effective zone of the circular element is shown to be much smaller than those of the other elements. The effects of Reynolds number and blockage ratio are investigated. A phenomenon of monoside vortex shedding is observed. The role of the element's bluffness is investigated and the suppression mechanism is discussed.

Numerical exploration of new features on three-dimensional transition of a cylinder wake

The three-dimensional transition of the wake flow behind a circular cylinder is studied in detail by direct numerical simulations using 3D incompressible N-S equations for Reynolds number ranging from 200 to 300. New features and vortex dynamics of the 3D transition of the wake are found and investigated. At Re = 200, the flow pattern is characterized by mode A instability. However, the spanwise characteristic length of the cylinder determines the transition features. Particularly for the specific spanwise characteristic length linear stable mode may dominate the wake in place of mode A and determine the spanwise phase difference of the primary vortices shedding. At Re = 250 and 300 it is found that the streamwise vortices evolve into a new type of mode - "dual vortex pair mode" downstream. The streamwise vortex structures switch among mode A, mode B and dual vortex pair mode from near wake to downstream wake. At Re = 250, an independent low frequency f(m) in addition to the vortex shedding frequency f(s) is identified. Frequency coupling between f(m) and f(s) occurs. These result in the irregularity of the temporal signals and become a key feature in the transition of the wake. Based on the formation analysis of the streamwise vorticity in the vicinity of cylinder, it is suggested that mode A is caused by the emergence of the spanwise velocity due to three dimensionality of the incoming flow past the cylinder. Energy distribution on various wave numbers and the frequency variation in the wake are also described.

Compressible laminar boundary-layer flows of a dusty gas over a semi-infinite flat-plate

The compressible laminar boundary-layer flows of a dilute gas-particle mixture over a semi-infinite flat plate are investigated analytically. The governing equations are presented in a general form where more reasonable relations for the two-phase interaction and the gas viscosity are included. The detailed flow structures of the gas and particle phases are given in three distinct regions : the large-slip region near the leading edge, the moderate-slip region and the small-slip region far downstream. The asymptotic solutions for the two limiting regions are obtained by using a seriesexpansion method. The finite-difference solutions along the whole length of the plate are obtained by using implicit four-point and six-point schemes. The results from these two methods are compared and very good agreement is achieved. The characteristic quantities of the boundary layer are calculated and the effects on the flow produced by the particles are discussed. It is found that in the case of laminar boundary-layer flows, the skin friction and wall heat-transfer are higher and the displacement thickness is lower than in the pure-gas case alone. The results indicate that the Stokes-interaction relation is reasonable qualitatively but not correct quantitatively and a relevant non-Stokes relation of the interaction between the two phases should be specified when the particle Reynolds number is higher than unity.

Vortex motion in the early stages of unsteady flow around a circular cylinder

In this paper, processes in the early stages of vortex motion and the development of flow structure behind an impulsively-started circular cylinder at high Reynolds number are investigated by combining the discrete vortex model with boundary layer theory, considering the separation of incoming flow boundary layer and rear shear layer in the recirculating flow region. The development of flow structure and vortex motion, particularly the formation and development of secondary vortex and a pair of secondary vortices and their effect on the flow field are calculated. The results clearly show that the flow structure and vortices motion went through a series of complicated processes before the symmetric main vortices change into asymmetric: development of main vortices induces secondary vortices; growth of the secondary vortices causes the main vortex sheets to break off and causes the symmetric main vortices to become “free” vortices, while a pair of secondary vortices is formed; then the vortex sheets, after breaking off, gradually extend downstream and the structure of a pair of secondary vortices becomes relaxed. These features of vortex motion look very much like the observed features in some available flow field visualizations. The action of the secondary vortices causes the main vortex sheets to break off and converts the main vortices into free vortices. This should be the immediate cause leading to the instability of the motion of the symmetric main vortices. The flow field structure such as the separation position of boundary layer and rear shear layer, the unsteady pressure distributions and the drag coefficient are calculated. Comparison with other results or experiments is also made.

后台阶下游床面冲刷过程数值模拟研究

以床面瞬时剪应力作为泥沙起动及输运的水动力机制,建立了结构物周围复杂流场下床面局部冲刷的数学模型。并应用大涡模拟方法对后台阶下游三维湍流流动进行数值模拟,得到台阶下游床面瞬时剪应力的分布规律。为了确定床面瞬时剪应力与泥沙上扬通量的关系,先应用数学模型对不同模型参数下,冲刷开始后5分钟时台阶下游床面形状进行试算。通过试算与实验结果的比较,确定床面瞬时剪应力与泥沙上扬通量关系中需要的模型参数。进一步对冲刷开始后30分钟内台阶下游床面演化规律进行模拟,模拟结果与实验结果相符合。

可燃气体中激波与障碍物作用在下游形成爆轰波的数值研究

对平面激波和单个矩形障碍物作用的过程进行了数值模拟,研究了反射产生的上行爆轰波在下游可燃气体中形成爆轰波的过程。数值结果表明,下游爆轰波形成过程主要有2种模式:爆轰波直接绕射和绕射波在上壁面反射,这和已有的实验结果是一致的。通过研究下游爆轰波的形成过程受入射激波马赫数、混合气体的压力和管道尺度的影响,分析了上游爆轰波向下游传播的波动力学过程,讨论了2种形成过程的作用规律和控制因素,阐明了下游爆轰波的形成规律。

Vortex dislocation in wake-type flow caused by local spanwise nonuniformity

DNS of spatiotemporal evolution of a wake-type flow is performed. In the incoming flow, a local spanwise nonuniformity in momentum defect is initially imposed. Results show that the spanwise nonuniformity leads to a series of symmetric twist vortex dislocation in downstream of the flow. Vortex line variations and substantial transition of vorticity from spanwise to the streamwise and vertical directions clearly feature the generation of a vortex dislocation and the real vortex linking in the dislocation. Dynamical process and the mechanism responsible for the vortex dislocation are described.

Effects of wall temperature on boundary layer stability over a blunt cone at Mach 7.99

Effects of wall temperature on stabilities of hypersonic boundary layer over a 7-degree half-cone-angle blunt cone are studied by using both direct numerical simulation (DNS) and linear stability theory (LST) analysis. Four isothermal wall cases with Tw/T0= 0.5, 0.7, 0.8 and 0.9, as well as an adiabatic wall case are considered. Results of both DNS and LST indicate that wall temperature has significant effects on the growth of disturbance waves. Cooling the surface accelerates unstable Mack II mode waves and decelerates the first mode (Tollmien–Schlichting mode) waves. LST results show that growth rate of the most unstable Mack II mode waves for the cases of cold wall Tw/T0=0.5 and 0.7 are about 45% and 25% larger than that for the adiabatic wall, respectively. Numerical results show that surface cooling modifies the profiles of rdut/dyn and temperature in the boundary layers, and thus changes the stability haracteristic of the boundary layers, and then effects on the growth of unstable waves. The results of DNS indicate that the disturbances with the frequency range from about 119.4 to 179.1 kHz, including the most unstable Mack modes, produce strong mode competition in the downstream region from about 11 to 100 nose radii. And adiabatic wall enhances the amplitudes of disturbance according to the results of DNS, although the LST indicates that the growth rate of the disturbance of cold wall is larger. That because the growth of the disturbance does not only depend on the development of the second unstable mode.

连续PCR扩增过程中异常现象的发生机理研究

聚合酶链式反应（Polymerase Chain Reaction，PCR）技术从其发明以来，因为其操作的简单方便和高效率而在生物学研究的各个领域得到了广泛的应用，包括序列扩增、序列的人工突变、疾病诊断、法医学鉴定、基因的表达分析等等。从PCR技术发明以来，如何提高反应的特异性和反应的效率一直是人们所共同关心的题目，为此也发展了相当数量的各种方法，如热启动PCR、降落PCR、巢式PCR以及在反应体系中添加一些有益的附属物等。而适合不同目的的PCR技术也得到了充分的发展，如多重PCR、反转录PCR、定量PCR、原位PCR、PCR突变、毛细管PCR技术等等。并且，包括随机引物扩增多态、扩增片段长度多态性、简单重复序列多态性、单核苷酸多态性等这些在PCR技术基础上发展而来的各种分子标记技术极大地方便了遗传分析和遗传图谱的构建等工作。在PCR技术发明了20年后的今天，提高PCR的反应性能、发展适合新领域的PCR技术和新的分子标记技术仍然是研究者关心的题目和努力的方向。 　　PCR实验中已经观察到多种异常现象，除了常见的扩增失败（没有产物）、扩增产物特异性不强（有非特异产物出现）、引物多聚体产物扩增、扩增效率低等现象以外，还包括PCR介导的重组、跳跃、复制滑动等等。阐明这些异常现象的发生机理和过程，避免或缓解这些异常现象在扩增过程中对目的产物扩增的影响，以及促进和利用一些特殊的异常PCR扩增都是PCR技术研究所关心的话题。各种研究工作中经常需要扩增一些长片段的序列，但是在进行长片段PCR时经常会发现扩增目标序列的长度是有限的、扩增效率比较低、扩增产物检测中有很强的背景弥散等现象;同时长片段PCR需要一些特殊的反应体系组成和反应条件。如何更加有效地实现更长序列的PCR扩增也是人们所关心的话题之一。 　　常见的PCR产物重复扩增（以上一轮扩增产物为模板进行新的PCR扩增）扩增轮数少，通常仅进行一次重复扩增;同时，在重复扩增中常使用的策略是使用巢式引物。而连续PCR扩增实验（用相同的引物以产物为模板进行多轮次的连续重复PCR扩增）从未见于文献报道。我们第一次系统地进行了连续PCR扩增实验;同时，在实验过程中我们观察到了一种新的PCR扩增异常现象——用不同来源的模板（病毒、细菌质粒或真核生物来源的DNA序列）进行连续PCR扩增不同长度的靶序列，经过有限次数的重复扩增后，最终都会导致扩增失败;这种扩增失败都表现为在常规琼脂糖电泳检测时特异产物条带的消失和不能泳动出点样孔之复杂异常产物的出现;这种扩增产生的异常产物能够被稳定地重复扩增。用λ和细菌质粒序列为模板连续扩增不同长度靶序列的结果表明：连续PCR扩增失败的时期具有扩增靶序列长度的依赖性，越长的靶序列在连续PCR中扩增失败的时期越早。 　　对不同连续PCR扩增的扩增过程观察表明扩增产物经历了一个从高效特异性扩增到低效率特异性扩增，再到扩增产生复杂异常产物的过程。对复杂异常产物的甲酰胺辅助变性处理和变性胶电泳（尿素变性聚丙烯酰胺胶电泳和NaOH碱变性琼脂糖电泳）检测表明扩增产生的复杂产物主要由连续分布的小于靶序列长度的具有相当程度多样性的非全长链组成。连续PCR产生的复杂产物在内部具有局部的双链区域和大量的单链区域及外部单链分支，能够被单链特异的S1核酸酶消化，但是不能被双链特异的限制性内切酶消化。用DNase I或限制性内切酶处理连续扩增早期产生特异扩增产物形成不同长度序列组成的混合物，或者直接用不同扩增反应产生的不同长度的核酸序列组成混合物，混合物在经历变性-复性后都表现出类似连续PCR失败所产生的异常产物电泳行为。这些证据都表明PCR扩增过程中形成的非全长链成分是导致这种异常现象的关键因素，多个不同长度的非全长链复性形成“杂种分子”（具有较大且不一致的分子量和复杂的分支结构），最终表现为常规琼脂糖电泳异常的复杂产物。同时，异常产物组成非全长链成分和全长链成分是其能够实现稳定重复扩增的基础。 　　实验结果表明：对于特定长度的靶序列而言，导致复杂异常出现的根本原因是连续PCR扩增体系中所经历的总PCR热循环数目（每一轮PCR扩增所使用的循环数目多，成功连续扩增的轮数就少）;而扩增体系中的引物浓度、DNA聚合酶用量的多少、扩增程序中时间参数等对此影响较小;巢式PCR和单引物-互补引物PCR的结果表明这些处理对于缓解或延迟异常产物的出现有一定的作用。人工处理（DNase I或限制性内切酶处理）完整模板双链形成的非全长链长产物，然后把非全长链长产物以不同比例同完整模板混合模拟连续扩增后期产物，这种人工混合模板表明连续PCR扩增中同源的非全长链成分对PCR扩增有严重的干扰作用，是导致复杂异常产物出现的直接原因。 　　已有的研究表明：PCR介导重组、长片段PCR难于操作有共同的产生基础——扩增过程中非全长链成分的产生和非全长链成分对后续扩增过程的干扰作用。这一点和导致连续PCR失败的原因是一致的。非全长链成分的出现是PCR扩增过程中不可避免的，其最初产生的可能来源有三个：模板的损伤（扩增前的模板损伤或扩增热循环过程中的损伤）、聚合酶的忠实性、以及聚合酶的进行性。根据聚合酶的特性而调整扩增程序中延伸时间的实验表明，聚合酶的进行性不是导致连续PCR扩增失败的最主要原因。这种非全长链成分产物从无到有且不依赖于体系中非全长链成分的过程我们称之为非全长链成分的初级合成;而已经存在的非全长链成分干扰后续合成形成非全长链成分的过程我们称之为非全长链成分的次级合成。非全长链成分的初级合成和次级合成共同导致了连续扩增的失败和异常产物的形成。 　　从已有的研究结果看，任何降低PCR扩增过程中非全长链成分产生的措施，特别是聚合酶忠实性的提高，都能缓解异常扩增产物的出现和利于长片段PCR操作。 　　

海南野生兰科植物保育生物学研究

兰科(Orchidaceae)是高等植物中第二大科，包括石斛属在内的许多种类均具有较高的观赏价值和独特的药用价值。过度采挖和商业利用及原生境丧失致使野生兰科植物资源受到了毁灭性的破坏，许多兰科植物种类日趋濒危。本研究以海南岛野生兰科植物为对象，从生物多样性、群落生态学、菌根体系的建立和重引入实施工程等方面开展工作，研究结果如下： 1）报道了中国兰科植物2个新种，1个中国新记录属，6个中国新记录种及11个海南新记录种。其中，2个新种为长筒天麻Gastrodia longitubularis和芳香白点兰Thrixspermum odoratum；1个中国新记录属为小囊兰属Micropera；6个中国新记录种，分别为疏花羊耳蒜Liparis sparsiflora、三色槌柱兰Malleola insectifera.、红花小囊兰Micropera poilanei、美丽云叶兰Nephelaphyllum pulchrum、腐生齿唇兰Odontochilus saprophyticu和白花曲唇兰Panisea albiflora；11个海南新记录种，分别为密花石豆兰Bulbophyllum odoratissimum、金唇兰Chrysoglossum ornatum、勐海隔距兰Cleisostoma menghaiense、束花石斛Dendrobium chrysanthum、无叶美冠兰Eulophia zollingeri、滇南翻唇兰Hetaeria rubens、心叶球柄兰Mischobulbum cordifolium、毛叶芋兰Nerviliaplicata、平卧曲唇兰Panisea cavalerei、云南曲唇兰Panisea yunnanensisi及台湾白点兰Thrixspermum formosanum。 2）海南琼北火山岩地区调查结果表明，该区有兰科植物10属14种，成斑块状分布；其区系成分单一，生活型不全面；附主植物种类多样，人工种植的经济作物为该地区兰科植物的主要附生宿主。生境特征分析结果显示，琼北火山岩地区独特的地质气候条件及植被类型是影响该地区兰科植物种类数量的主要因素。人工干扰所致的植被破坏、生境破碎化是该地区兰科植物多样性丧失的主导因素。 3）以我国兰科海南特有种-华石斛（Dendrobium sinense TangetWang）为研究对象，采用双向指示种分析法（TWINSPAN）和除趋势对应分析法（DCA）对海南霸王岭国家级自然保护区华石斛附生群落进行了分类和排序。结果表明：TWINSPAN分类结果将22个样方划分为2个植被类型、5个群系、7个群丛组；并分别对各群丛组的生境特点、植被树种组成作了论述。DCA二维排序图较好地验证了TWINSPAN的分类结果，并客观地反映了群落的生境特征。随着海拔逐渐升高，华石斛附生群落植被类型从山地常绿林逐渐过渡到高山苔藓矮林，群落中的优势种由喜高温耐干旱的热带、亚热带树种逐渐演变为耐低温喜湿的亚热带、温带树种。华石斛在主要群落类型群系间、群丛组内的数量、密度和空间分布格局表现出较大差异，以具通风、凉爽和高湿环境的高山苔藓矮林附生数量最多。附生群落类型的多样性反映了华石斛对多样生境的适应性。华石斛附生群落植被类型群丛组的划分，进一步细化和丰富了海南热带森林植被类型山地常绿林和高山矮林的划分，有助于了解华石斛自然居群的生境特点，为进一步开展华石斛的保育工作提供了科学依据。 4）兰科菌根菌对兰花种子萌发和原球茎生长起了重要作用，在成年阶段也扮演着重要角色。为了解东南亚特有种五唇兰的菌根形态和菌根真菌的动态侵染过程，对不同生境下两种变异类型（叶背绿色和叶背紫色）的成年植株菌根分三个时间段采样并进行显微观察。结果表明，成年的五唇兰植株与菌根真菌有着密切关系，菌丝的侵染、定殖和消解程度随着生长发育变化而变化；在营养生长阶段和生殖生长阶段都发现菌丝结，这两个阶段的菌丝特征有所不同；而在休眠阶段的菌根中观察不到菌丝结；不同生境和不同变异类型的菌根结构无明显区别。 5)利用种子原地共生萌发技术，通过种子预处理及播种等技术的不断完善，获得了大量种子萌发原球茎；初步揭示了华石斛种子萌发原球茎规律。结果表明：10%NaOH浸泡10min预处理能显著提高华石斛种子萌发率；原生境人工播种明显提高了萌发的原球茎数及宿主原球茎产生率；但不同附生宿主间存在差异，自然萌发原球茎数非常低。播种后2.5个月左右种子开始萌发，4.5个月左右达到高峰，在6.5个月原球茎数基本趋于稳定。 6)华石斛菌根化苗重引入实验结果表明，实验室筛选的有益真菌与原生境下筛选有益真菌存在一定差异。菌根化苗重引入野外生境，3个月后以12号华石斛菌根化苗生长最好，3、9号华石斛菌根化苗生长较正常，而7、10号两种华石斛菌根化苗叶片受损或死亡，无菌组培苗基本已死亡；华石斛菌根化苗成活率在50%以上，而对照几乎全部死亡；从菌根化苗长势评价，3、9、12号3个内生真菌华石斛菌根化苗生长较好，为真正有益共生真菌。

转基因植物生产生物可降解塑料PHB的研究-phbA功能鉴定、多价表达载体的构建及转基因植物的获得

聚-β-羟基链烷酸（PHA）是许多微生物作为碳源、能源的一类贮藏性聚酯，具有广泛的应用价值。该聚酯可被微生物完全降解且有与塑料相似的性质，因而研究并提高PHA在植物中的合成为解决环境污染提供了新的解决途径。 聚-β-羟基于酸酯（PHB）是研究的最早、研究的最清楚的一种PHA。用聚合酶链式反应扩增并克隆了真养产碱杆菌（Alcaligenes eutrophus）中合成PHB的一个关键酶——3-酮硫裂解酶基因phbA。DNA序列分析表明所克隆的基因与国外报道序列同源性很高，只有一个碱基对的区别。为了检测该基因的功能及导肽的定位效率，构建了带有导肽基因的组成型表达载体，由根癌农杆菌介导转化烟草（Nicotiana tabacum cv. Wisconsin 38）得到转基因植株。蛋白质电泳结果表明导肽可以将外源蛋白定位于质体，phbA基因能翻译成相应大小的蛋白。酶活性分析证实了转基因烟草中phbA编码的3-酮硫裂解酶可以催化乙酰-CoA合成乙酰乙酰-CoA。 将携有导肽序列的phbC（编码PHB合酶）和phbB（编码乙酰乙酰-CoA还原酶）连入pBIB-HYG得到组成型表达载体pZCB，用冻融法转入根癌农杆菌，介导转化烟草。烟草为已获得的具有卡那霉素抗性整合并表达phbA的转基因烟草。通过二次转化将携有潮霉素抗性的phbB基因和phbC基因导入已整合phbA的烟草，各基因均由质体导肽控制，最后得到整合PHB合成的三个酶基因的转基因烟草。转基因烟草经PCR、PCR-Southern检测，初步确定整合phbB和phbC烟草植株。以气相色谱初步分析，转基因烟草中PHB的含量可达鲜重的0.233%。 结果表明phbB和phbC基因可以在真核表达系统中编码相应的蛋白。通过色素分析、荧光动力学等手段分析了PHB在叶绿体中的累积对其功能的影响。 为了提高底物乙酰-CoA的供应能力及减少惰性聚酯对植物体的伤害，分离了种子特异性启动子和质体导肽序列，利用忆经克隆的合成PHB的三个关键酶基因，通过一系列DNA重组，分别构建了含有种子特异性启动子的嵌合phbC、phbB的二价表达载体pSCB及嵌合phbC、phbA、phbB的三价表达载体pSCAB，并由导肽将基因表达产物定位于质体。经根癌农杆菌介导转化油菜（Brassica napus L.) H165，获得转基因油菜植株，并进行了PCR、Southern blot及RT-PCR-DNA杂交等分检测。结果表明，三基因已经分别整合到相应的转基因油菜中，并已在转录水平表达。同时转化了油菜不育系、恢复系和保持系，获得批量转化株，并移入温室栽培。

多糖衍生物的钒络合物及其应用

本发明公开了一种多糖衍生物的钒络合物的制备方法，首先将NaOH碱化了的多糖与一氯乙酸在40-80℃下反应3-4小时，合成多糖衍生物；然后将上述多糖衍生物与钒溶液在室温下反应12-24小时，得到多糖衍生物的钒络合物。该多糖衍生物的钒络合物应用在治疗糖尿病药物中。本发明采用多糖的衍生物为原料，在水性介质中，中性条件下与VOSO4反应，经过过滤、干燥、粉碎，得到钒与多糖衍生物的络合物。本发明中，络合物的制备完全在室温条件下进行，工艺简单，易于实现工业化生产。

GPIb is involved in platelet aggregation induced by mucetin, a snake C-type lectin protein from Chinese habu (Trimeresurus mucrosquamatus) venom

Mucetin (Trimeresurus mucrosquamatus venom activator, TMVA) is a potent platelet activator purified from Chinese habu (Trimeresurus mucrosquamatus) venom. It belongs to the snake venom heterodimeric C-type lectin family and exists in several multimeric forms. We now show that binding to platelet glycoprotein (GP) lb is involved in mucetin-induced platelet aggregation. Antibodies against GPIb as well as the GPIb-blocking C-type lectin echicetin inhibited mucetin-induced platelet aggregation. Binding of GPIb was confirmed by affinity chromatography and Western blotting. Antibodies against GPVI inhibited convulxin- but not mucetin-induced aggregation. Signalling by mucetin involved rapid tyrosine phosphorylation of a number of proteins including Syk, Src, LAT and PLCgamma2. Mucetininduced phosphorylation of the Fcgamma chain of platelet was greatly promoted by inhibition of alpha(llb)beta(3) by the peptidomimetic EMD 132338, suggesting that phosphatases downstream Of alpha(llb)beta(3) activation are involved in dephosphorylation of Fcgamma. Unlike other multimeric snake C-type lectins that act via GPIb and only agglutinate platelets, mucetin activates alpha(llb)beta(3). Inhibition Of alpha(llb)beta(3) strongly reduced the aggregation response to mucetin, indicating that activation Of alpha(llb)beta(3) and binding of fibrinogen are involved in mucetin-induced platelet aggregation. Apyrase and aspirin also inhibit platelet aggregation induced by mucetin, suggesting that ADP and thromboxaneA(2) are involved in autocrine feedback. Sequence and structural comparison with closely related members of this protein family point to features that may be responsible for the functional differences.