第四章--细胞质膜

1、第四章 细胞质膜,第一节 细胞质膜的结构模型 第二节 细胞膜基本特征与功能 第三节 膜骨架,第一节 细胞质膜的结构模型,一、细胞膜(cell membrane)：又称质膜(plasma membrane)，是围绕在细胞表面，由脂质和蛋白质组成的生物膜。由于它的存在，使细胞成为一个相对独立的结构，并通过它使细胞与环境之间能够进行物质、能量的交换及信息的传递。 对于细胞膜结构和功能的认识，有一个由表及里、不断深入的有趣历程，产生了几个结构模型。,（一）三明治模型 1. 细胞膜脂双层概念的提出 1925年两位荷兰科学家E.Gorter和F.Grendel分离纯化了红细胞，从一定数量的红细胞中抽提脂类

2、，按Langmuir的方法进行展层，并比较展层后的脂单层的面积和根据体积所推算的总面积， Gorter和Grendel发现提取的脂铺展后所测的面积同实际测量的红细胞的表面积之比约为1.82.21，为了解释这一结果，他们提出红细胞膜的基本结构是脂双层结构的概念。,Langmuir 的研究工作,2. 随后，人们发现质膜的表面张力比油-水界面的表面张力低得多，因此推测质膜中含有蛋白质成分，并提出“蛋白质-脂质-蛋白质”的三明治质膜结构模型。3. 1959年，Robertson 用超薄切片技术获得了清晰的细胞膜照片，显示暗-明-暗三层结构，这就为三明治模型提供了证据，并提出了单位膜模型，认为所有的生物

3、膜都是由“蛋白质-脂质-蛋白质”的单位膜构成。单位膜模型是三明治模型发展。,（二）流动镶嵌模型 (fluid mosaic model)。 S. J. Singer Glc = glucose, GalNAc = N-acetylgalactos-amine; these three sugars are uncharged.,3. 胆固醇（cholesterol）,主要存在真核细胞膜上，但植物细胞膜中含量较少，其功能是：调节膜的流动性，增加膜的稳定性，降低水溶性物质的通透性。胆固醇的结构如下图：,Cholesterol in a lipid bilayer. Schematic drawin

4、g of a cholesterol molecule interacting with two phospholipid molecules in one leaflet of a lipid bilayer.,（二）膜脂的运动方式,1.沿膜平面的侧向运动（基本运动方式）,其扩散速率为10-8cm2/s； 2.脂分子围绕轴心的自旋运动； 3.脂分子尾部的摆动； 4.双层脂分子之间的翻转运动，发生频率还不到脂分子侧向交换频率的1010。但在内质网膜上,新合成的磷脂分子翻转运动发生频率很高。,Figure 10-6. Phospholipid mobility. The types of mov

5、ement possible for phospholipid molecules in a lipid bilayer.,（三）脂质体,脂质体(lipsome)是根据磷脂分子可在水相中形成稳定的脂双层的趋势而制备的人工膜。,(a)水溶液中的磷脂分子团；(b)球形脂质体；(c)平面脂质体膜；(d）用于疾病治疗的脂质体的示意图,脂质体的应用,研究膜脂与膜蛋白及其生物学性质； 脂质体中裹入DNA可用于基因转移； 在临床治疗中,脂质体作为药物或酶等载体,二、 膜蛋白 膜蛋白是膜功能的主要体现者。核基因组编码的蛋白质中30%左右的为膜蛋白。根据与膜脂的结合方式以及在膜中的位置的不同，膜蛋白分为内在蛋白

6、（integral protein）、外周蛋白（peripheral protein）和脂锚定蛋白（lipid-anchored protein）。,蛋白与膜的结合方式 、整合蛋白；、脂锚定蛋白；、外周蛋白,（一）内在蛋白（integral proteins） 内在蛋白又称为整合蛋白，以不同程度嵌入脂双层的内部，有的为全跨膜蛋白（tansmembrane proteins）。膜蛋白为两性分子。它与膜结合非常紧密，只有用去垢剂（detergent）才能从膜上洗涤下来，常用SDS和Triton-X100。 内在蛋白的跨膜结构域形成亲水通道有两种形式，一是由多个螺旋组成亲水通道；二是由折叠组成亲水通

7、道。,内在蛋白与脂膜的结合方式 膜蛋白的跨膜结构域与脂双层分子的疏水核心的相互作用。 跨膜结构域两端带正电荷的aa残基与磷脂分子带负电的极 性头形成离子键；或带负电的氨基酸残基通过Ca2+、Mg2+等 阳离子与带负电的磷脂极性头相互作用。 膜蛋白在细胞质基质一侧的半胱氨酸残基上共价结合脂肪 酸分子,插入脂双层之间, 还有少数蛋白与糖脂共价结合。,A typical single-pass transmembrane protein. Note that the polypeptide chain traverses the lipid bilayer as a right-handed a h

8、elix and that the oligosaccharide chains and disulfide bonds are all on the noncytosolic surface of the membrane. Disulfide bonds do not form between the sulphydryl groups in the cytoplasmic domain of the protein because the reducing environment in the cytosol maintains these groups in their reduced

9、 (-SH) form.,跨膜结构域与膜脂作用的方式,20个左右疏水氨基酸残基形成螺旋，外部疏水侧链通过范德华力与脂双层分子脂肪酸相互作用； 某些螺旋具有极性和非极性两种侧链，形成特异极性分子的跨膜通道； 某些跨膜蛋白由1012个氨基酸形成折叠结构，反向折叠结构可以形成非特异性的跨膜通道，可允许相对分子量小于10K的分子自由通过。,A segment of a transmembrane polypeptide chain crossing the lipid bilayer as an a helix. Only the a-carbon backbone of the polypeptid

10、e chain is shown, with the hydrophobic amino acids in green and yellow. (J. Deisenhofer et al., Nature 318:618-624 and H. Michel et al., EMBO J. 5:1149-1158),The three-dimensional structure of a bacteriorhodopsin molecule. The polypeptide chain crosses the lipid bilayer as seven a helices. The locat

11、ion of the chromophore and the probable pathway taken by protons during the light-activated pumping cycle are shown. When activated by a photon, the chromophore is thought to pass an H+ to the side chain of aspartic acid 85. Subsequently, three other H+ transfers are thought to complete the cyclefro

12、m aspartic acid 85 to the extra-cellular space, from aspartic acid 96 to the chromophore, and from the cytosol to aspartic acid 96. (R. Henderson et al. J. Mol. Biol.213:899-929),细菌紫膜质,Figure. The three-dimensional structure of the photosynthetic reaction center of the bacterium Rhodopseudomonas vir

13、idis. The structure was determined by x-ray diffraction analysis of crystals of this transmembrane protein complex. The complex consists of four subunits, L, M, H, and a cytochrome. The L and M subunits form the core of the reaction center, and each contains five a helices that span the lipid bilaye

14、r. The locations of the various electron carrier coenzymes are shown in black. (Adapted from a drawing by J. Richardson based on data from J. Deisenhofer et al., Nature 318:618-624),The three-dimensional structure of a porin trimer of Rhodobacter capsulatus determined by x-ray crystallography. (A) E

15、ach monomer consists of a 16-stranded antiparallel b barrel that forms a transmembrane water-filled channel. (B) The monomers tightly associate to form trimers, which have three separate channels for the diffusion of small solutes through the bacterial outer membrane. A long loop of polypeptide chai

16、n (shown in red), which connects two b strands, protrudes into the lumen of each channel, narrowing it to a cross-section of 0.6 x 1 nm. (Adapted from M.S. Weiss et al., FEBS Lett.280: 379-382),Unlike most other integral membrane proteins, which are essentially alpha-helical, the porin consists pred

17、ominantly of beta-sheet structures.The integral membrane protein porin from R.capsulatus consists of three tightly associated 16-stranded beta-barrels that give rise to three distinct diffusion channels for small solutes through the outer membrane. The X-ray structure of this porin has revealed deta

18、ils of its shape, the residue distributions within the pore and at the membrane-facing surface, and the location of the calcium sites.,（二）外周蛋白（peripheral protein） 外周蛋白又称为外在蛋白（extrinsic protein），为水溶性的，分布在细胞膜的表面，靠离子键或其它较弱的键与膜表面的蛋白质分子或脂分子的亲水部分结合，因此只要改变溶液的离子强度甚至提高温度就可以从膜上分离下来。,（三）脂锚定蛋白（lipid-anchored pr

19、otein） 脂锚定蛋白通过磷脂或脂肪酸锚定，共价结合。分两类，一类是糖磷脂酰肌醇(GPI)连接的蛋白，GPI位于细胞膜的外小叶，用磷脂酶C处理能释放出结合的蛋白。许多细胞表面的受体、酶、细胞粘附分子和引起羊瘙痒病的PrPC就是这类蛋白。另一类脂锚定蛋白与插入质膜内小叶的长碳氢链结合。,A detergent micelle in water, shown in cross-section. Because they have both polar and nonpolar ends, detergent molecules are amphipathic.,（四）研究膜蛋白的常用试剂：去垢剂

20、(detergent),Solubilizing membrane proteins with a mild detergent. The detergent disrupts the lipid bilayer and brings the proteins into solution as protein-lipid-detergent complexes. The phospholipids in the membrane are also solubilized by the detergent.,The structures of two commonly used detergen

21、ts. Sodium dodecyl sulfate (SDS) is an anionic detergent, and Triton X-100 is a nonionic detergent. The hydrophobic portion of each detergent is shown in green, and the hydrophilic portion is shown in blue. Note that the bracketed portion of Triton X-100 is repeated about eight times.,The use of mil

22、d detergents for solubilizing, purifying, and reconstituting functional membrane protein systems. In this example functional Na+-K+ ATPase molecules are purified and incorporated into phospholipid vesicles. The Na+-K+ ATPase is an ion pump that is present in the plasma membrane of most animal cells;

23、 it uses the energy of ATP hydrolysis to pump Na+ out of the cell and K+ in.,第二节 生物膜基本特征与功能,一、膜的流动性,（一）膜脂的流动性 主要指脂分子的侧向运动 影响因素： 脂肪酸链的饱和度：脂肪酸链所含双键越多越不饱和，使膜流动性增加。 脂肪酸链的链长：长链脂肪酸相变温度高，膜流动性降低。 胆固醇：胆固醇的含量增加会降低膜的流动性。 其他因素：温度、酸碱度、离子强度等。,Experiment demonstrating the mixing of plasma membrane proteins on mous

24、e-human hybrid cells. The mouse and human proteins are initially confined to their own halves of the newly formed heterocaryon plasma membrane, but they intermix with time. The two antibodies used to visualize the proteins can be distinguished in a fluorescence microscope because fluorescein is gree

25、n whereas rhodamine is red. (Based on observations of L.D. Frye and M. Edidin, J. Cell Sci. 7:319-335),（二）膜蛋白的流动性,Antibody-induced patching and capping of a cell-surface protein on a white blood cell. The bivalent antibodies cross-link the protein molecules to which they bind. This causes them to cl

26、uster into large patches, which are actively swept to the tail end of the cell to form a cap. The centrosome, which governs the head-tail polarity of the cell, is shown in orange.,光脱色恢复技术 (fluorescence recovery after photobleaching, FRAP),对细胞某一部位进行光照脱色，对此处荧光恢复的速度进行检测，从而得知膜流动性的特征。,Diagram of an epith

27、elial cell showing how a plasma membrane protein is restricted to a particular domain of the membrane. Protein A (in the apical membrane) and protein B (in the basal and lateral membranes) can diffuse laterally in their own domains but are prevented from entering the other domain, at least partly by

28、 the specialized cell junction called a tight junction. Lipid molecules in the outer (noncytoplasmic) monolayer of the plasma membrane are likewise unable to diffuse between the two domains; lipids in the inner (cytoplasmic) monolayer, however, are able to do so.,Three domains in the plasma membrane

29、 of guinea pig sperm defined with monoclonal antibodies. A guinea pig sperm is shown schematically in (A), while each of the three pairs of micrographs shown in (B), (C), and (D) shows cell-surface immunofluorescence staining with a different monoclonal antibody (on the right) next to a phase-contra

30、st micrograph (on the left) of the same cell. The antibody shown in (B) labels only the anterior head, that in (C) only the posterior head, whereas that in (D) labels only the tail. (Courtesy of Selena Carroll and Diana Myles.),Four ways in which the lateral mobility of specific plasma membrane prot

31、eins can be restricted. The proteins can self-assemble into large aggregates (such as bacteriorhodopsin in the purple membrane of Halobacterium) (A); they can be tethered by interactions with assemblies of macromolecules outside (B) or inside (C) the cell; or they can interact with proteins on the s

32、urface of another cell (D).,质膜的流动性是保证其正常功能的必要条件。例如跨膜物质运输、细胞信息传递、细胞识别、细胞免疫、细胞分化以及激素的作用等等都与膜的流动性密切相关。当膜的流动性低于一定的阈值时，许多酶的活动和跨膜运输将停止，反之如果流动性过高，又会造成膜的溶解。,膜流动性的意义,二、 膜的不对称性,（一）细胞膜各部分的名称 质膜内外两层的组分和功能的差异，称为膜的不对称性。 样品经冰冻断裂处理后，细胞膜可从脂双层中央断开，各断面命名为：ES，细胞外表面（extrocytoplasmic surface）；EF，细胞外小页断面（extrocytoplasmic

33、face）；PS，原生质表面（protoplasmic surface）；PF，原生质小页断面（protoplasmic face） 。,（二）膜脂的不对称性,The asymmetrical distribution of phospholipids and glycolipids in the lipid bilayer of human red blood cells. glycolipids are drawn with hexagonal polar head groups (blue).,（三）膜蛋白的不对称性 每种膜蛋白分子在细胞膜上都具有特定的方向性和分布的区域性。如各种激素的

34、受体具有极性，细胞色素C位于线粒体内膜M侧。 质膜上的糖蛋白其糖残基均分布在质膜的ES面。 生物膜的特征及其生物学功能主要是由膜蛋白来决定的，膜蛋白的不对称性是生物膜完成复杂的在时间与空间上有序的各种生理功能的保证。,三、细胞膜的功能, 为细胞的生命活动提供相对稳定的内环境。 选择性的物质运输,包括代谢底物的输入与代谢 产物的排除,其中伴随着能量的传递。 提供细胞识别位点,并完成细胞内外信息跨膜传递。 为多种酶提供结合位点,使酶促反应高效而有序地进行。 介导细胞与细胞、细胞与基质之间的连接。 参与形成具有不同功能的细胞表面特化结构。 质膜参与形成具有不同功能的细胞表面特化结构。,第三节 膜骨架

35、,细胞质膜常常与膜下结构相互联系、协同作用，并形成细胞表面的一些特化结构以完成特定的功能。这些结构包括膜骨架、鞭毛和纤毛、微绒毛及细胞的变形足等，分别与细胞形态的维持、细胞运动、细胞的物质交换等功能有关。,一、膜骨架,指细胞质膜下与膜蛋白相连的由纤维蛋白组成的网架结构,它参与维持细胞质膜的形状并协助质膜完成多种生理功能。,研究膜骨架的模式细胞-哺乳动物的红细胞。,二、红细胞的生物学特性 哺乳动物成熟的红细胞没有细胞核和内膜系统，所以它是最简单、最易研究的生物膜。 红细胞质膜既具有很好的弹性又具有较高的强度，是因为其膜下有一个膜骨架结构。 红细胞被低渗破裂后，其中的血红蛋白等可溶性成分被释放出去

36、，剩下的结构仍然保持原来的基本形状和大小，这种结构称为血影（blood ghost）.,三、红细胞质膜蛋白及膜骨架,SDS polyacrylamide-gel electrophoresis pattern of the proteins in the human red blood cell membrane. The gel in (A) is stained with Coomassie blue. The positions of some of the major proteins in the gel are indicated in the drawing in (B); gl

37、ycophorin is shown in red to distinguish it from band 3. Other bands in the gel are omitted from the drawing. The large amount of carbohydrate in glycophorin molecules slows their migration so that they run almost as slowly as the much larger band 3 molecules. (A, courtesy of Ted Steck.),1.血影蛋白 (spe

38、ctrin),是红细胞膜骨架的主要成份,但不是红细胞膜的成份,约占膜提取蛋白的30%。血影蛋白属红细胞的膜下蛋白，这种蛋白是一种长的、可伸缩的纤维状蛋白,长约100 nm,由两条相似和亚基构成。两个亚基链呈反向平行排列, 扭曲成麻花状，形成异二聚体, 两个异二聚体头-头连接成200 nm长的四聚体。5个或6个四聚体的尾端一起连接于短的肌动蛋白纤维并通过非共价键与带4.1蛋白结合,而带4.1 蛋白又通过非共价键与跨膜蛋白带3蛋白或血型糖蛋白的细胞质面结合, 形成“连接复合物”。这些血影蛋白在整个细胞膜的细胞质面下面形成可变形的网架结构,以维持红细胞的双凹圆盘形状。,Spectrin molecu

39、les from human red blood cells. The protein is shown schematically in (A) and in electron micrographs in (B). Each spectrin heterodimer consists of two antiparallel, loosely intertwined, flexible polypeptide chains called and these are attached noncovalently to each other at multiple points, includi

40、ng both ends. The phosphorylated head end, where two dimers associate to form a tetramer, is on the left. Both the a and b chains are composed largely of repeating domains 106 amino acids long. In (B) the spectrin molecules have been shadowed with platinum. (D.W. Speicher and V.T. Marchesi, Nature 3

41、11:177-180; B, D.M. Shotton et al., J. Mol. Biol. 131:303-329),The spectrin-based cytoskeleton on the cytoplasmic side of the human red blood cell membrane. The structure is shown schematically in (A) and in an electron micrograph in (B). The arrangement shown in (A) has been deduced mainly from studies on the interactions of purified proteins in vitro. Spectrin dimers associate head-to-hea