Phase transitions and heterogeneity in lipid bilayers

The optical reflectivity of several well-characterized lipid bilayer systems has been correlated with calorimetric studies of the membrane components. There is a large increase in mean membrane thickness when a bilayer is cooled below the transition temperature of the membrane lipid. Similar studies on membranes generated from a mixture of two lipids possessing different degrees of unsaturation suggest that between the characteristic transition temperatures of the two lipids, the bilayer contains clusters of gel and liquid crystalline lipid which coexist within the plane of the membrane.     

Membrane structure: some general principles

The arrangement of lipids and some proteins in the erythrocyte membrane has been discussed. The conclusions from this are listed here as a set of general guidelines for the structure of membranes of higher organisms: some of these rules may be wrong. But at this stage it seems useful to sharpen our thoughts in this way and thereby focus attention on various specific points. 1) The basis of a membrane is a lipid bilayer with (i) choline phospholipids and glycolipids in the external half and (ii) amino (and possibly some choline) phospholipids in the cytoplasmic half. There is effectively no lipid exchange across the bilayer (unless enzymatically catalyzed) (68). 2) Some proteins extend across the bilayer. Where this is so, they will in general have carbohydrate on their surface remote from the cytoplasm. This carbohydrate may prevent the protein diffusing out of the membrane into the cytoplasm; it acts as a lock on the protein. 3) Just as lipids do not flip-flop, proteins do not rotate across the membrane. Lateral motion or rotation of lipids and proteins in the plane of the bilayer may be expected. 4) Most membrane protein is associated with the inner, cytoplasmic, urface of the membrane. Proteins are not usually associated exclusively with the outer half of the lipid bilayer. 5) Membrane proteins are a special class of cytoplasmic proteins, not of secreted proteins.     

A mutant of Arabidopsis lacking a chloroplast-specific lipid

In order to investigate the functional significance of membrane lipid unsaturation, we have isolated a series of mutants of Arabidopsis thaliana which are deficient in particular membrane fatty acids. The first of these mutants completely lacks Delta3-trans-hexadecenoate, an acyl group that until now has been thought to play an important role in the structure or function of thylakoid membranes in photosynthetic eukaryotes. The apparent absence of any marked physiological effect of the mutation illustrates the potential of this approach to the analysis of membrane structure and function.     

Surface-induced x-ray reflection visualization of membrane orientation and fusion into multibilayers

The fusion of lipid membranes at the air-water interface has been detected with the use of x-ray reflection as a high-resolution, surface-sensitive technique. The rate of this fusion for dimyristoylphosphatidylcholine (DMPC) bilayers is the highest at 29 degrees C, which coincides with the chain-melting phase-transition temperature for the top membrane layers. After 6 hours of incubation a stack of at least ten surface-ordered membrane bilayers in equilibrium with the bulk vesicle suspension is formed. Such fusion is thus surface-catalyzed but not restricted to the first surface layer. The process involves partial membrane dehydration near the solution surface which decreases toward the bulk.     

Phase separation of lipid membranes analyzed with high-resolution secondary ion mass spectrometry

Lateral variations in membrane composition are postulated to play a central role in many cellular events, but it has been difficult to probe membrane composition and organization on length scales of tens to hundreds of nanometers. We present a high-resolution imaging secondary ion mass spectrometry technique to reveal the lipid distribution within a phase-separated membrane with a lateral resolution of approximately 100 nanometers. Quantitative information about the chemical composition within small lipid domains was obtained with the use of isotopic labels to identify each molecular species. Composition variations were detected within some domains.     

Protein conformations in the plasma membrane

Infrared spectroscopy and optical rotatory dispersion have been used to test theories of structure of membrane protein. No evidence has been found to support the view that adjacent to the lipid there is a monolayer of protein in the beta-conformation. The extracted protein appears to be a fairly typical globular protein with a low a-helical content.     

Physiological regulation of the immunological synapse by agrin

T cell activation is dependent on both a primary signal delivered through the T cell receptor and a secondary costimulatory signal mediated by coreceptors. Although controversial, costimulation is thought to act through the specific redistribution and clustering of membrane and intracellular kinase-rich lipid raft microdomains at the contact site between T cells and antigen-presenting cells. This site has been termed the immunological synapse. Endogenous mediators of raft clustering in lymphocytes have not been identified, although they are essential for T cell activation. We now demonstrate that agrin, an aggregating protein crucial for formation of the neuromuscular junction, is also expressed in lymphocytes and is important in reorganization of membrane lipid microdomains and setting the threshold for T cell signaling. Our data show that agrin induces the aggregation of signaling proteins and the creation of signaling domains in both immune and nervous systems through a common lipid raft pathway.     

Restricted lateral diffusion of PH-20, a PI-anchored sperm membrane protein

The rate of lateral diffusion of integral membrane proteins is constrained in cells, but the constraining factors for most membrane proteins have not been defined. PH-20, a sperm surface protein involved in sperm-egg adhesion, was shown to be anchored in the plasma membrane by attachment to the lipid phosphatidylinositol and to have a diffusion rate that is highly restricted on testicular sperm, being more than a thousand times slower than lipid diffusion. These results support the hypothesis that lateral mobility of a membrane protein can be regulated exclusively by interactions of its ectodomain.     

柔嫩艾美耳球虫敏感株与抗药株子孢子膜脂流动性的检测

建立了应用荧光探针1,6-二苯基-1,3,5-己三烯(DPH)测定柔嫩艾美耳球虫子孢子膜脂流动性的方法。对柔嫩艾美耳球虫(Eimeria tenella)的敏感株与各种抗药株共13个球虫株的子孢子膜脂流动性进行测定。结果表明,敏感株的膜脂流动性大,抗药株膜脂流动性小(P0.01,呈极显著相关)。由此可见,虫株对药物的敏感性可能与膜脂流动性有关,膜脂流动性的检测结果可作为判别柔嫩艾美耳球虫对离子载体类抗球虫药物敏感性或抗药性的指标之一。     

Dynamic structure of membranes by deuterium NMR

Progress in our understanding of the dynamic structure of membrane lipids and proteins has recently been made possible by the advent of high-field "solid-state" nuclear magnetic resonance spectroscopic studies of specifically deuterium-labeled systems. Major features of lipid and protein dynamics have been deduced.     

The direction of membrane lipid flow in locomoting polymorphonuclear leukocytes

The objective of this study was to determine the direction of membrane lipid flow in locomoting cells. The plasma membrane of human polymorphonuclear leukocytes was stained with a fluorescent lipid analog dihexadecanoyl indocarbocyanine. A line was photobleached on the cell surface perpendicular to the direction of cell motion. Low-light-level fluorescence microscopy and digital image-processing techniques were used to analyze a series of images taken at short intervals after photobleaching. The bleached line remained visible for about 5 seconds before being erased by diffusional recovery. Examination of fluorescence intensity profiles allowed a comparison to be made between the velocities of line and cell movement. Results indicate that the bleached line moves forward with the same velocity as the cell during locomotion, refuting the retrograde lipid flow model of locomotion. Instead, the plasma membrane lipid appears to move forward according to either the unit movement of membrane or the tank track model of locomotion.     

Partitioning of lipid-modified monomeric GFPs into membrane microdomains of live cells

Many proteins associated with the plasma membrane are known to partition into submicroscopic sphingolipid- and cholesterol-rich domains called lipid rafts, but the determinants dictating this segregation of proteins in the membrane are poorly understood. We suppressed the tendency of Aequorea fluorescent proteins to dimerize and targeted these variants to the plasma membrane using several different types of lipid anchors. Fluorescence resonance energy transfer measurements in living cells revealed that acyl but not prenyl modifications promote clustering in lipid rafts. Thus the nature of the lipid anchor on a protein is sufficient to determine submicroscopic localization within the plasma membrane.     

Penetration of a Single Domain of Bacillus thuringiensis Cry1Ie-Domain I to a Lipid Membrane In vitro

Domain I of the activated Crystal protein from Bacillus thuringiensis has a seven α-helix bundle structure, which is responsible for membrane channel formation in its insecticidal mechanism. Cry1Ie is toxic to Asian corn borer, Ostrinia furnacalis (Guenée), and plays important roles in insect biological control. The domain I from Cry1Ie has been expressed and purified in its normal conformation, as embedded in the full length homologous toxin structure. The membrane insertion ability of this single domain was compared with the full length homologous toxin using a monolayer insertion experiment. The results indicated that the Cry1Ie-domain I had the ability to insert into the lipid monolayer, and this ability is greater than that of the IE648 toxin. However, the state of insertion is not stable and remains for only a short period of time. The Cry1Ie-domain I plays no role in receptor binding as it had a nonspecific binding with the brush border membrane vesicles of the Asian corn borer.     

膜类脂组成和脂肪酸含量对种子耐干性的影响

种子的耐干性与膜脂的组成和脂肪酸含量有一定关系,与质膜中脂肪酸的不饱和度关系可能更为密切。分别选用代表淀粉类、蛋白质类和油料类种子的玉米（Zea maysL.）、榆树（Ulmus pumilaL.）和草红花（Carthamus tinctoriusL.）种子为试材,运用薄层层析法和气相色谱法研究了3种类型种子膜类脂的组成,通过分析不同品种间生物膜类脂的组成差异对细胞膜结构和功能的影响,进而揭示不同类型种子耐干性的关系。结果表明：3类种子中榆树种子的极性脂组成最丰富且含量最多,玉米种子次之,草红花种子最少,说明膜类脂在蛋白质类种子生物膜中起的作用大于油料类和淀粉类种子;不同类型种子膜类脂的组成明显不同,脂肪酸含量变化显著,表明不同的膜类脂组分在保持膜系统的完整性和生物膜正常的代谢过程中所起的作用不同,油料种子草红花具有特殊的耐干机理。     

采后鸭梨衰老与膜脂过氧化的关系

鸭梨采后衰老时，硬度下降，膜脂过氧化产物丙二醛含量增加，质膜透性增大．低温、钙处理可抑制果实的膜脂过氧化作用，降低质膜透性，延缓了果实衰老．果实过氧化氢酶活性变化幅度较大，其活性后期高于前期，且不同温度下变化曲线不同，果实硬度与丙二醛含量呈负相关．由此认为，膜脂过氧化作用是鸭梨衰老的一个重要因素．     

Leader peptidase of Escherichia coli: critical role of a small domain in membrane assembly

Leader peptidase spans the Escherichia coli plasma membrane with its amino-terminal domain facing the cytoplasm and its carboxyl terminus facing the periplasm. It is made without a cleavable leader sequence. The three apolar domains near the amino terminus of the peptidase are candidates for internal "signal sequences" and they anchor the protein to the lipid bilayer. Oligonucleotide-directed deletion was used to show that only the second domain has an essential function in membrane assembly. While this second apolar domain is crucial for membrane assembly, its continued function when disrupted by arginine suggests that its apolar character per se is not its only important feature.     

水分胁迫下水杨酸对油松幼苗叶片膜脂过氧化作用的影响

采用盆栽PEG处理的方法研究了水分胁迫条件下水杨酸对油松幼苗膜脂过氧化作用的影响.结果表明,水分胁迫导致油松幼苗SOD和CAT活性降低,O-2积累,由此引起膜脂过氧化,MDA含量增高,质膜相对透性增大.在水分胁迫期间外加水杨酸(SA)处理可提高SOD和CAT活性,降低O-和MDA含量.这表明在水分胁迫条件下SA能够降低膜脂过氧化作用,对膜脂具有保护作用.     

衰老小麦叶片膜脂的变化及6－BA的作用

以小麦叶片为试材．研究了衰老过程中膜脂的变化。结果表明．随着衰老膜磷脂含量下降．种类发生变化．脂氧合酶活性增加．脂质过氧化产物ＭＤＡ累积．膜透性增加。６－ＢＡ减缓磷脂分解，降低脂氧合酶活性从而降低膜脂过氧化，维持膜完整性。     

脂滴的生物物理学和细胞生物学研究

最早对脂滴的研究可以追溯到19 世纪,Richard Altmann 和E.B. Wilson 都描述了细胞 中的脂肪滴,并推测了它们的起源(Altmann, 1890;威尔逊,1896)。早期,脂滴的高衍射特性为其 鉴定提供了便利的光学条件。20 世纪初,由于它们被认为是大多数细胞的组成部分,这种细胞 器被称为脂质体。然而,在20 世纪60 年代末,人造脂质体被发明出来,并很快取代了这个名 字。从那时起,这种细胞器被有过许多名称,包括脂滴、脂体、脂体、脂滴和脂质体。在植物中, 它们通常被称为油体。随着这一领域的迅速发展,它似乎已经定名为“脂滴”。几十年来,除了 形态学研究,脂滴的其他方面几乎没有受到关注。1991 年,脂肪细胞中与脂滴相关的磷酸蛋白 perilipin 的发现,引起人们对这种细胞器的新关注。从那时起,关于脂滴的论文数量急剧增加, 关于脂滴的功能也逐渐被重视起来。     

烤烟烘烤过程中叶片膜脂过氧化特性的研究

采用温湿度自动控制电热烤烟箱，研究了烘烤过程中烟叶膜脂过氧化及其内源保护酶活性的变化。结果表明，随烘烤时间延长，叶内超氧物歧化酶（ＳＯＤ）和过氧化物酶（ＰＯＤ）活性下降，丙二醛（ＭＤＡ）含量增加。烘烤４８ｈ和７２ｈ后，两种酶先后失活，ＭＤＡ积累达最大值，证明烟叶烘烤过程中叶片膜脂过氧化水平不断提高。不同处理以高温快烤烟叶膜脂过氧化水平最高，低温慢烤最低。而且变黄温度对烟叶膜脂过氧化的作用比定色期不同升温处理的作用大。