Chapter 10: Membrane Structure. Membrane Structure Membrane Function ► Plasma membrane defines...

Chapter 10: Chapter 10: Membrane StructureMembrane Structure

Membrane StructureMembrane Structure

Membrane FunctionMembrane Function► Plasma membrane defines cell and maintains differences Plasma membrane defines cell and maintains differences

btwn cytosol and extracellular environmentbtwn cytosol and extracellular environment► Defines individual organellesDefines individual organelles► Establish ion gradientsEstablish ion gradients► Contain proteins act as sensors of external signals allow cell Contain proteins act as sensors of external signals allow cell

to adapt to changing environmentto adapt to changing environment

Membrane StructureMembrane Structure

General Membrane Structure

►Thin film of lipid and protein held together by noncovalent interactions

►lipid bilayer serves as basic fluid structure; impermeable barrier

►Proteins mediate all other functions of membrane

The Lipid BilayerThe Lipid Bilayer

Lipids of MembraneLipids of Membrane► 50% of total membrane mass, 5 x 1050% of total membrane mass, 5 x 1066/um/um22

► AmphipathicAmphipathic► Self-sealingSelf-sealing► Phosphlipids, cholestrol, glycolipidsPhosphlipids, cholestrol, glycolipids► Phospholipids= most abundant lipid in membranePhospholipids= most abundant lipid in membrane► Spontaneously aggregate to bury hydrophobic tailSpontaneously aggregate to bury hydrophobic tail

The Lipid BilayerThe Lipid BilayerFluidity of Lipid BilayerFluidity of Lipid Bilayer► Rapid lateral diffusion of phospholipids 10Rapid lateral diffusion of phospholipids 10-8-8cm/seccm/sec► Change places 10Change places 1077 times/sec times/sec► Rarely flip-flopRarely flip-flop► Fluidity depends on composition (phospholipid and cholesterol) Fluidity depends on composition (phospholipid and cholesterol)

and temperatureand temperature► Phase transition= the temperature at which there is a change of Phase transition= the temperature at which there is a change of

state from liquid to solidstate from liquid to solid


Fluidity and length and saturation of FA hydrocarbon chainsFluidity and length and saturation of FA hydrocarbon chains► Short hydrocarbon chain lengths fluidityShort hydrocarbon chain lengths fluidity► Double bonds fluidityDouble bonds fluidity


Fluidity and cholesterol contentFluidity and cholesterol content

Cholesterol fluidityCholesterol fluidity Provides mechanical stabilityProvides mechanical stability 1 cholesterol/phospholipid in eucaryotes1 cholesterol/phospholipid in eucaryotes No cholesterol in procaryotes; mechanical No cholesterol in procaryotes; mechanical

stabililty imparted by cell wallstabililty imparted by cell wall


Major Membrane Phospholipids Major Membrane Phospholipids


Lipid RaftsLipid Rafts► Microdomains enriched in sphingolipids, cholesterol and Microdomains enriched in sphingolipids, cholesterol and

membrane proteinsmembrane proteins► Long saturated FA chain of sphingolipids = attractive forces that Long saturated FA chain of sphingolipids = attractive forces that

hold adjacent molecules togetherhold adjacent molecules together► Thicker than other parts of bilayerThicker than other parts of bilayer► able to accommodate membrane proteins concentrating for able to accommodate membrane proteins concentrating for

transport or to enable proteins to function togethertransport or to enable proteins to function together


Membrane asymmetryMembrane asymmetry► Phospholipid distribution: Phospholipid distribution:

phosphatidylcholine and sphingomyelin confined to outer monolayer phosphatidylcholine and sphingomyelin confined to outer monolayer phosphatidylethanolamine and phosphatidylserine are on inner phosphatidylethanolamine and phosphatidylserine are on inner

monolayer monolayer ► Charge Charge ► Proteins Proteins ► Impt to function (ie, apoptosis and translocation of phosphatidylserine) Impt to function (ie, apoptosis and translocation of phosphatidylserine) ► glycolipids glycolipids


Glycolipids Glycolipids ► Sugar containing lipid molecules w/ Sugar containing lipid molecules w/

most extreme assemmetry in most extreme assemmetry in distributiondistribution

► Found exclusively on Found exclusively on noncytoplasmic monolayernoncytoplasmic monolayer

► On surface of all plasma membranesOn surface of all plasma membranes► gangliosides= most complex, sialic gangliosides= most complex, sialic

acid containing oligosaccharides, acid containing oligosaccharides, net negative chg, most abundant in net negative chg, most abundant in pm of nerve cellspm of nerve cells

► Function- protection, cell Function- protection, cell recognition, transmission of recognition, transmission of electrical impulseselectrical impulses

Membrane ProteinsMembrane Proteins

Proteins Carry Out Specific Functions of Membrane

►Give ea type of membrane characteristic functional properties

►Amts and types of proteins highly variable

►By mass proteins represent 50% lipids 50%; lipids small thus 1 protein/ 50 lipid molecules

►Associate w/ membrane in various ways depending on function: transmembrane, integral, peripheral

Membrane ProteinsMembrane ProteinsTransmembrane Proteins Typically Cross as Alpha HelixTransmembrane Proteins Typically Cross as Alpha Helix► Unique orientation reflection syn and insertion in ER and functionUnique orientation reflection syn and insertion in ER and function► Membrane spanning doamin comprised of nonpolar aa 20-30Membrane spanning doamin comprised of nonpolar aa 20-30► Alpha helix is predominate conformation but beta shees form Alpha helix is predominate conformation but beta shees form

closed beta barrels that can span membrane as wellclosed beta barrels that can span membrane as well► Can predict membrane spanning regions via hydropathy plotCan predict membrane spanning regions via hydropathy plot


Beta Barrels form Transmembrane ChannelsBeta Barrels form Transmembrane Channels► Tend to be more rigidTend to be more rigid► Comprised of 8-22 strandsComprised of 8-22 strands► Some pore forming proteins generating water-filled channels for Some pore forming proteins generating water-filled channels for

select hydrophilic solutesselect hydrophilic solutes► Polar side chains line channel on inside; nonpolar side chains project Polar side chains line channel on inside; nonpolar side chains project

out interact w/hydrophobic core of lipid bilayerout interact w/hydrophobic core of lipid bilayer


Many Membrane Proteins are Glycolslated

►Majority of transmembrane proteins in animal cells are glycolsylated

►Sugars added in lumen of ER and golgi

►Oligosaccharide chains always present on noncytosolic side


Membrane Proteins can be Solubilized and Purified in Detergents

Distrupt hydrophobic interactions

Bind to hydrophobic regions of membrane and displace lipid molecules

SDS, triton


Red Blood Cells

Model system for studying membranes

Available in lg numbers

Easy to isolate uncontaminated from other cell types

No nucleus or internal organelles

Can prepare ghosts and in-side-out vesicles


Ghosts and InSide-Out RBCsGhosts and InSide-Out RBCsGhosts= empty RBCs Ghosts= empty RBCs

prepared by placing cells in soln of low salt to cause water to move into and lyse cellsprepared by placing cells in soln of low salt to cause water to move into and lyse cellscan reseal or be studied while leakycan reseal or be studied while leaky

Inside-Out VesiclesInside-Out Vesicles


Use of Sealed and Unsealed RBC ghostsUse of Sealed and Unsealed RBC ghosts Demonstrated some proteins extend across lipid bilayerDemonstrated some proteins extend across lipid bilayer Enabled sidedness of proteins to be determinedEnabled sidedness of proteins to be determined

Label intact sealed ghosts and inside-out vesicles w/ water Label intact sealed ghosts and inside-out vesicles w/ water solutble label that cannot penetrate lipid bilayer– perform SDS solutble label that cannot penetrate lipid bilayer– perform SDS PAGEPAGE

Exposed internal or external surface to proteolytic enzymes that Exposed internal or external surface to proteolytic enzymes that are membrane impremeant (transmembrane protein will be are membrane impremeant (transmembrane protein will be partially digested from both sidespartially digested from both sides

Labeled antibodies Labeled antibodies


Studies of RBC Plasma Membrane Proteins by SDS Studies of RBC Plasma Membrane Proteins by SDS PAGEPAGE

15 major protein bands btwn 15,000-250,000 daltons15 major protein bands btwn 15,000-250,000 daltons 3 most prominent comprose more than 65% protein mass; ea 3 most prominent comprose more than 65% protein mass; ea

arranged differently in membranearranged differently in membrane SpectrinSpectrin GlycophorinGlycophorin Band 3Band 3


Spectrin= Cytoskeletal Protein Assoc. w/ Cytosolic Side of Spectrin= Cytoskeletal Protein Assoc. w/ Cytosolic Side of MembraneMembrane

Most proteins are peripheral and assoc w/ cytosolic sideMost proteins are peripheral and assoc w/ cytosolic side Spectrin most abundantSpectrin most abundant Long, thin, flexible rod 100 nm length, constitutes 25% protein massLong, thin, flexible rod 100 nm length, constitutes 25% protein mass 2.5 x 102.5 x 1055 copies/cell copies/cell Principle component of cytoskeleton underlying RBC, maintains Principle component of cytoskeleton underlying RBC, maintains

structural integrity and biconcave shapestructural integrity and biconcave shape Heterodimer formed from 2 lg structurally similar subunits that assoc Heterodimer formed from 2 lg structurally similar subunits that assoc

head to head to form 200 nm long tetramerhead to head to form 200 nm long tetramer Tail ends of 4-5 tetramers linked by binidng short actin filaments and Tail ends of 4-5 tetramers linked by binidng short actin filaments and

other cytoskeletal components forming meshwork other cytoskeletal components forming meshwork Abnormalities in spectrin results in anemia and spherical shaped RBCs Abnormalities in spectrin results in anemia and spherical shaped RBCs

that are fragilethat are fragile Ankyrin binds spectrin and band 3 to membraneAnkyrin binds spectrin and band 3 to membrane


Spectrin


GlycophorinGlycophorin sm singlepass glycoprotein of 131 aasm singlepass glycoprotein of 131 aa Most of mass on external surface of Most of mass on external surface of

membranemembrane 100 sugar residues on 16 diff side chains 100 sugar residues on 16 diff side chains

which account for 60% protein masswhich account for 60% protein mass >90% sialic acid most of negative chg >90% sialic acid most of negative chg

on surface of RBCon surface of RBC 1 million molec/cell1 million molec/cell Function unknownFunction unknown


Band 3Band 3 Multipass membrane proteinMultipass membrane protein Catalyzes couple transport of anionsCatalyzes couple transport of anions 930 aa thought to extend across bilayer 12X930 aa thought to extend across bilayer 12X Allows HCOAllows HCO33 to cross membrane in exchg for Cl to cross membrane in exchg for Cl--

increasing amt of COincreasing amt of CO22 delivered to lungs delivered to lungs


Bacteriorhodopsin is a Proton Pump

Halobacterium salinarum “purple membrane”

7 transmembrane helices of 25 aa

Retinal light absorbing chromophore linked to lys side chain

Light excites chromophore causing conf chg which results in transfer of H+ from inside to outside cell

Electrochemical gradient used to syn ATP

Can pump up to 100 H+/sec


Membrane Proteins Often Function as Lg ComplexesMembrane Proteins Often Function as Lg Complexes


Membrane Proteins Diffuse in Plane of Membrane Proteins Diffuse in Plane of

Membrane but Do Not Flip-FlopMembrane but Do Not Flip-Flop


Techniques for Measuring the Lateral Techniques for Measuring the Lateral

Diffusion of Membrane ProteinsDiffusion of Membrane Proteins


Confining Proteins and Lipids to Confining Proteins and Lipids to Specific Specific

Regions of the MembraneRegions of the Membrane


Cell surface Coated with Sugar Residues

Glycocalyx= CHO rich region on cell surface

Proteoglycans= long polysaccharide chains linked to protein core

Oligosahharides also linked to proteoglycans

Possbile functions: protection, receptors, electrical conductivity

Chapter 10: Membrane Structure. Membrane Structure Membrane Function ► Plasma membrane defines...

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