BMB 462 Lecture 3 Outline of Last Lecture I. Introduction to Membrane lipidsa. Basic Structureb. Basic propertiesII. Classes of Membrane lipidsa. Glycerophospholipidsb. Galactolipids/Sulfolipidsc. Sphingolipidsd. Sterolse. IsoprenoidsIII. Lipase activityIV. Lipid SignalingOutline of Current Lecture I. Experimentation with LipidsII. Membrane FunctionIII. Membrane Structure & Compositiona. Common featuresb. Lipid Compositionc. Fluid Mosaic Modeld. Protein CompositionIV. Introduction to Hydropathy PlotsCurrent LectureConcepts to remembers from previous courses/lectures:- GLC and HPLC procedures- Amino Acid structures and properties (polar amino acids vs. hydrophobic amino acids)I. Experimentation with Lipidsa. Lipids are poorly soluble in H2O so when separating out lipids, you need them to end in an organic solventi. Proteins, nucleic acids, etc, would end up in the aqueous solution.b. Isolation of Lipids Using Organic Solvents:These notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.i. Lipids are hydrophobic so they partition into hydrophobic organic solventsii. Neutral lipids are extracted with nonpolar solvents (like chloroform)iii. Membrane lipids are extracted with more polar organic solvents (i.e. methanol) that weaken both the hydrophobic and electrostatic interactionsc. Separation via Absorption Chromatography or Thin-Layer Chromatography (TLC)i. Once you’ve separated the lipids from the rest of the cell components, you need to separate the different types of lipidsii. Differences in solubility in nonpolar solvents or polarity of different lipids allows separationiii. Polar lipids bind to the silica while neutral lipids move into the organic solventiv. Increasing the polarity of the solvent removes increasingly polar lipidsv. Chromatography is based on partitioning between a nonpolar mobile phase (which gets increasingly more polar) and a polar stationary phased. Determination of chain length and saturation - by GLC or HPLC (mass spectrometer protocols)e. Determination of fatty acid position – by enzymatic degradationf. Determination of double bond positions – mass spec.- Beginning of unit on Membrane Function, Structure, and Dynamics -II. Membrane Functiona. Membranes regulate what crosses into/out of the cell i. They are semi-permeable; transport proteins aid in movement of moleculesb. They set boundaries and compartments within the cell c. They can organize complex reactions allowing communication and energy generation i. Receptor proteins in membranes are important for detecting signalsii. The 2D surface of membranes increases molecular interactions and increases efficiencyd. Exocytosis/Endocytosis possible because membranes can break and resealIII. Membrane Structure & Compositiona. Common featuresi. Sheet-like structures 5-8nm thickii. Amphipathic, arranged in lipid bilayeriii. Membrane components are not covalently bound but are linked by noncovalent interactions that allow the membrane to remain fluidiv. Consist of polar lipids and proteins; these often have carbohydrates attachedv. Asymmetricvi. Electrically polarizedb. Lipid Compositioni. Membranes are composed of different types and ratios of proteins, phospholipids, sterols, and other lipids, according to what tissue they are a part of and what molecules they need.1. e.g. Why would you need more proteins in a bacterial membrane than in a human myelin sheath?E. coli has to have more of a barrier to protect itself from the environment. Most of E. coli’s energy comes from the membrane (it comes from mitochondria in the myelin sheath; the membrane in myelin is typically just insulation)ii. Most common lipids in (almost all) membranes: sphingolipids, phosphatidylcholine, phosphatidylethanolamineiii. A lot of proteins in the membranes means the membrane has a lot of activity/functionsiv. Cells have a way of targeting and maintaining lipids in the membrane; the lipid membrane has important functions according to associated tissue1. Basic membrane functions: protection, structure, etc – membranes also have tissue-specific/needs-specific functions2. There is a functional importance to the side of the membrane a phospholipid is ona. e.g. if Phosphatidylserine is in the inner leaflet, everything in the cell is ok. If the cell is dying and degrading, it will signal this to other cells by switching PS to the outer leaflet. This signals surrounding cells that the cell is going to degrade, which brings the other cells in to help with apoptosis or get clear of the area.v. In the bilayer of the membrane, the inner (facing cytosol) and outer (facing external environment) leaflets, the composition is asymmetrical1. Components in the outer leaflet are different from those found in the inner leafleta. Due to random flip-flopping of membrane components, it would seem logical that there would eventually be an evendistribution of all molecules. This doesn’t happen, though, because the cell exerts effort to maintain asymmetry.c. Fluid Mosaic Modeli. Components are lipids, proteins (peripheral, integral, lipid-linked), carbohydrates (linked to protein or lipid, on the outer face)ii. Movement accounts for the “fluid” portion; the components are not static or rigid but move laterally (along the same leaflet) or transversely (across leaflets)1. Lipids are not bound covalently; they rely on hydrogen bonding, Van der Waals interactions, etc, to stay together.2. Lateral movement happens frequently and easily; transverse movement is more difficult for the cell (because the polar head has to move through the hydrophobic area to get to the other side) and happens much less commonly.iii. Asymmetry accounts for the “Mosaic” portion of the model. d. Protein Compositioni. There are 3 types of membrane proteins: peripheral, integral, lipid-linked1. Peripheral Proteinsa. These are only found on the polar surface of the membraneb. They are held in place by electrostatic interactions and hydrogen bonds (they don’t penetrate into the hydrophobic core; they interact mostly with the polar heads)c. Peripheral proteins can be removed by causing changes in salt concentration or pHi. These are very mild treatments that don’t largely impact the membraneii. Adding detergent or an organic solvent would disrupt the membrane and make it too difficult to separate the peripheral protein from other membrane proteins/components2. Integral
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