MEMBRANES This text is divided int seven majr sectins: Overview f membranes The chemical cmpnents f membranes Membrane structure Membranes and cmpartmentalizatin Membrane receptrs Sme receptrs invlve secnd messengers Insulin and grwth factr receptrs Overview f membranes. The cell is nt an amrphus sack f cmpnents, but a cmplex structure filled with rganelles. Examples include: 1. endplasmic reticulum 2. mitchndria 3. nucleus Membranes are nt passive barriers. 1. They cntrl the structures and envirnments f the cmpartments they define, and thereby the metablism f these cmpartments. 2. The membrane itself is a metablic cmpartment with unique functins. Membranes are dynamic. 1. They can mve. 2. Their cmpnents are cntinuusly synthesized and degraded. 3. The primary event in cell death (e.g. mycardial infarctin) may be damage t the cell membrane, leading ultimately t cell death. Preview: In the fllwing sectins we will lk at membranes frm the perspectives f 1. chemical cmpnents 2. structure 3. functin The chemical cmpnents f membranes
General cmpsitin. 1. The cmpnents Lipid -- chlesterl, phsphlipid and sphinglipid Prteins Carbhydrate -- as glycprtein 2. Differences in cmpsitin amng membranes (e.g. myelin vs. inner mitchndrial membrane) Illustrate the variability f membrane structure. This is due t the differences in functin. Example: Mitchndrial inner membrane has high amunts f functinal electrn transprt system prteins. Plasma membrane, with fewer functins (mainly in transprt), has less prtein. Membranes with similar functin (i.e. frm the same rganelle) are similar acrss species lines, but membranes with different functin (i.e. frm different rganelles) may differ strikingly within a species. Distributin f lipids in membranes 1. There can be large membrane t membrane differences in lipids (cmpare phsphglycerides and chlesterl in plasma membrane vs. inner mitchndrial membrane). 2. There can be large differences within classes f lipids (cmpare the cardilipin f the inner mitchndrial membrane t ther membranes). 3. There are als patterns f differences amng the fatty acyl grups f the lipids f varius membranes The reasns fr these variatins are nt knwn.
The prteins f membranes. 1. Classificatin f membrane prteins is peratinal. Definitin f "peratinal classificatin": based upn hw a thing respnds t a specified treatment (r peratin), rather than upn the intrinsic nature f the thing. There are tw kinds f membrane prteins Extrinsic (r peripheral): These may be remved frm the membrane, r slubilized, by mild treatment, such as shaking with a dilute salt slutin. Extrinsic prteins are ften thught f as being lsely assciated with the membrane surface. Intrinsic (r integral): These cannt be remved frm the membrane withut treatment that destrys the membrane structure, such as disslving it with detergent. Intrinsic prteins are ften pictured as being deeply imbedded in the membrane r transfixing it. The bilgical activity f sme prteins depends n whether r nt they are assciated with membrane. 2. Rles f membrane prteins. Catalytic: enzymes Receptrs fr signals such as hrmnes. Binding f the hrmne t the prtein transmits the signal. Transprt Structural. The bicncave discidal shape f the erythrcyte is due in part t the membrane prtein. Carbhydrates f membranes are present attached t prtein r lipid as glycprtein r glyclipid. 1. Typical sugars in glycprteins and glyclipids include glucse, galactse, mannse, fucse and the N-acetylated sugars like N-acetylglucsamine, N-acetylgalactsamine and N-acetylneuraminic acid (sialic acid). 2. Membrane sugars seem t be invlved in identificatin and recgnitin. Membrane structure
The amphipathic prperties f the phsphglycerides and sphinglipids are due t their structures. 1. The hydrphilic head bears electric charges cntributed by the phsphate and by sme f the bases. These charges are respnsible fr the hydrphilicity. Nte that n lipid bears a psitive charge. They are all negative r neutral. Thus membranes are negatively charged. 2. The lng hydrcarbn chains f the acyl grups are hydrphbic, and tend t exclude water. 3. Phsphlipids in an aqueus medium spntaneusly aggregate int rderly arrays. Micelles: rderly arrays f mlecular dimensins. Nte the hydrphilic heads riented utward, and the hydrphbic acyl grups riented inward. Micelles are imprtant in lipid digestin; in the intestine they assist the bdy in assimilating lipids. Lipid bilayers can als frm. Lipsmes are structures related t micelles, but they are bilayers, with an internal cmpartment. Thus there are three regins assciated with lipsmes: The exterir The membrane itself The inside. Lipsmes can be made with specific substances disslved in the interir cmpartment. These may serve as mdes f delivery f these substances. 4. The prperties f phsphlipids determine the kinds f mvement they can underg in a bilayer. Mdes f mvement that maintain the hydrphilic head in cntact with the aqueus surrundings and the acyl grups in the interir are permitted. Rtatin Lateral diffusin Flexing f the acyl chains Transverse mvement frm side t side f the bilayer (flip-flp) is relatively slw, and is nt cnsidered t ccur significantly.
Membranes are currently pictured accrding t the fluid msaic mdel. 1. A lipid bilayer cmpsed f phsphlipid and chlesterl 2. Prteins. Integral prteins are shwn; peripheral prteins may be lsely attached t the surface. The difficulty with which flip-flp mvement f membrane cmpnents ccurs relates t the sidedness f membranes. Membrane surfaces have asymmetry -- different characteristics n the tw sides. 1. There are differences in lipid cmpsitin between the sides f a membrane. The mechanism fr generating this sidedness is unknwn. 2. Membranes als shw sidedness with respect t prtein cmpsitin Different catalytic prteins (enzymes) appear n the tw sides f membranes. Carbhydrate is mstly n the uter surface f cell membranes. It is typically attached t the prtin f membrane prteins that sticks ut. 3. The erythrcyte membrane prvides a gd mdel f membrane sidedness. Sme prteins (ankyrin, spectrin) are assciated with the inner surface f the membrane. Other prteins transfix the membrane (glycphrin), r lp back and frth frm side t side (band 3 prtein). Nte that there is carbhydrate n the exterir prtin f glycphrin and band 3. Membrane fluidity -- accrding t the fluid msaic mdel, prteins and lipids diffuse in the membrane. Membranes separate and maintain the chemical envirnments f the tw sides f the membrane. Intrductin: there are in gradients acrss the mammalian plasma membrane. Here is a cmparisn f the mean cncentratin f selected ins utside and inside a typical mammalian cell, giving the in, the cncentratin in the extracellular fluid, the intracellular fluid and the difference betwen the tw. Na + 140 mm 10 mm 14-fld K + 4 mm 140 mm 35-fld Ca ++ 2.5 mm 0.1 micrm 25,000-fld Cl - 100 mm 4 mm 25-fld Cell membranes maintain these gradients by preventing in flux active transprt f ins frm side t side f the plasma membrane.
Sme substances can crss membranes by passive (simple) diffusin. 1. Types f mlecules that can crss membranes by diffusin: Water and small lipphilic rganic cmpunds can crss. Large mlecules (e.g. prteins) and charged cmpunds d nt crss. 2. Directin relative t the cncentratin gradient: mvement is DOWN the cncentratin gradient ONLY (higher cncentratin t lwer cncentratin). 3. Rate f diffusin depends n charge n the mlecule -- electric charge prevents mvement. size -- smaller mlecules mve faster than larger mlecules. lipid slubility -- mre highly lipid-sluble mlecules mve faster. the cncentratin gradient -- the greater the cncentratin difference acrss the membrane, the faster the diffusin. 4. Directin relative t the membrane: mlecules may crss the membrane in either directin, depending nly n the directin f the gradient. Prtein channels transprt specific ins. 1. In channels exist fr Na +, K + and Ca ++ mvement. These channels are specific fr a given inic species. 2. Channels cnsist f prtein, which frms a gate that pens and clses under the cntrl f the membrane ptential. 3. In mvement thrugh channels is always dwn the cncentratin gradient. Transprt f mlecules acrss membranes by carriers (mediated transprt). 1. A carrier must be able t perfrm fur functins in rder t transprt a substance. Recgnitin -- t specifically bind the substance that is t be transprted. Translcatin -- mvement frm ne side f the membrane t the ther. Release -- n the ther side f the membrane
Recvery -- return f the carrier t its riginal cnditin s it can g thrugh anther cycle f transprt. 2. Terminlgy: Carriers are als variusly called "prters,""prting systems,""translcases,""transprt systems" and "pumps." 3. Carriers resemble enzymes in sme f their prperties. They are NOT enzymes, as they d NOT catalyze chemical reactins. They are enzyme-like in the fllwing ways. They are specific. They have dissciatin cnstants fr the transprted substances which are analgus t Km f enzymes. Transprt can be inhibited by specific inhibitrs. They exhibit saturatin, like enzymes d. Diffusin, in cntrast, is nt saturable, and its rate increases with increasing cncentratin. 4. A general mdel fr transprt is that the carrier is a prtein which changes cnfrmatin during the transprt prcess. 5. Smetimes carriers mve mre than ne mlecule simultaneusly. Nmenclature: Uniprt: a single mlecule mves in ne directin. Symprt: tw mlecules mve simultaneusly in the same directin. Antiprt: Tw mlecules mve simultaneusly in ppsite directins. Passive mediated transprt, r facilitated diffusin. 1. The characteristics f a carrier perating by passive mediated transprt. Faster than simple diffusin Mvement is dwn the cncentratin gradient nly (like diffusin) N energy input is required -- the necessary energy is supplied by the gradient. The carrier exhibits specificity fr the structure f the transprted substance saturatin kinetics specific inhibitability 2. Examples f passive mediated transprt. Glucse transprt in many cells. A uniprt system Can be demnstrated by the fact that adding substances with structures that resemble the structure f glucse can inhibit glucse transprt specifically. It is specific fr glucse. The K m fr glucse is 6.2 mm (a value in the neighbrhd f the bld cncentratin f glucse, 5.5 mm) The K m fr fructse is 2000 mm The transprt prcess invlves attachment f glucse utside the cell. Cnfrmatinal change f the carrier prtein. Release f the glucse inside the cell. There is n need t change K m fr glucse, since the glucse cncentratin in the cell is very lw. Chlride-bicarbnate transprt in the erythrcyte membrane. This is catalyzed by the band 3 prtein seen previusly. An antiprt system: bth ins MUST mve in ppsite directins simultaneusly. The system is reversible, and can wrk in either directin. Mvement is driven by the cncentratin gradient.
Active mediated transprt invlves transprt against a cncentratin gradient, and requires energy. 1. There are tw surces f energy fr active transprt. ATP hydrlysis may be used directly. The energy f the Na + gradient may be used in a symprt mechanism. The energy f the Na + ging dwn its gradient drives the mvement f the ther substance. But since the Na + gradient is maintained by ATP hydrlysis, ATP is the indirect surce f energy fr this prcess. 2. The characteristics f a carrier perating by active transprt. Can mve substances against (up) a cncentratin gradient. Requires energy. Is unidirectinal The carrier exhibits specificity fr the structure f the transprted substance saturatin kinetics specific inhibitability 3. Hw can the substance be released frm the carrier int a higher cncentratin than the cncentratin at which it bund in the first place? The affinity f the translcase fr the substance must decrease, presumably by a cnfrmatinal change f the translcase. This prcess may require energy in the frm f ATP. 4. Examples f active mediated transprt. Ca ++ transprt is a uniprt system, using ATP hydrlysis t drive the Ca ++ mvement. There are tw Ca ++ translcases f imprtance. In the sarcplasmic reticulum, imprtant in muscle cntractin. A different enzyme with similar activity in the plasma membrane. The Na + -K + pump (r Na + -K + ATPase). An antiprt system. Imprtance: present in the plasma membrane f every cell, where its rle is t maintain the Na + and K + gradients. Stichimetry: 3 Na + are mved ut f the cell and 2 K+ are mved in fr every ATP hydrlyzed. Specificity: Abslutely specific fr Na +, but it can substitute fr the K +. The structure f the Na + -K + pump is a tetramer f tw types f subunits, alpha 2 beta 2. The beta-subunit is a glycprtein, with the carbhydrate n the external surface f the membrane. The Na + -K + ATPase is specifically inhibited by the uabain, a carditnic sterid. Ouabain sensitivity is, in fact, a specific marker fr the Na + -K + ATPase. The prpsed mechanism f the Na + -K + ATPase shws the rle f ATP in effecting the cnfrmatinal change. Na + attaches n the inside f the cell membrane. The prtein cnfrmatin changes due t phsphrylatin f the prtein by ATP, and the affinity f the prtein fr Na + decreases. Na + leaves. K + frm the utside binds. K + dephsphrylates the enzyme. The cnfrmatin nw returns t the riginal state. K + nw dissciates.
Na + linked glucse transprt is fund in intestinal mucsal cells. It is a symprt system; glucse is transprted against its gradient by Na + flwing dwn its gradient. Bth are transprted int the cell frm the intestinal lumen. Na + is required; ne Na + is carried with each glucse. The Na + gradient is essential; it is maintained by the Na + -K + ATPase. Na + linked transprt f amin acids, als fund in intestinal mucsal cells, wrks similarly. There are at least six enzymes f different specificity that emply this mechanism. Their specificity is as fllws. Shrt neutral amin acids: ala, ser, thr. Lng r armatic neutral amin acids: phe, tyr, met, val, leu, ile. Basic amin acids and cystine: lys, arg, cys-cys. Acidic amin acids: glu, asp Imin acids: pr and hypr Beta-amin acids: beta-alanine, taurine. Membrane receptrs Cell-cell cmmunicatin is by chemical messenger. 1. There are fur types f signals. Nerve transmissin Hrmne release Muscle cntractin Grwth stimulatin 2. There are fur types f messenger mlecules. sterids small rganic mlecules peptides prteins 3. The messenger may interact with the cell in either f tw ways. Entry int the cell by diffusin thrugh the cell membrane (the sterid hrmnes d this). Large mlecules r charged nes bind t a receptr n the plasma membrane. 4. The events assciated with cmmunicatin via these mlecules may include the fllwing. Primary interactin f the messenger with the cell (binding by a receptr). A secndary event, frmatin f a secnd messenger. (this is nt always fund). The cellular respnse (sme metablic event). Terminatin (remval f the secnd messenger). Messenger mlecules which diffuse int the cell -- example: sterid hrmnes. 1. Sterids are lipid sluble, and can diffuse thrugh the plasma membrane. 2. Cells which are sensitive t sterid hrmnes have specific receptr prteins in the cytsl r nucleus which bind the sterid. 3. The receptr-hrmne cmplex then smehw causes changes in the cell's metablism, typically by affecting transcriptin r translatin. 4. The mechanism f terminatin is unclear, but invlves breakdwn f the hrmne. Plasma membrane receptrs.
1. Membrane receptrs bind specific messenger mlecules n the exterir surface f the cell. Either f tw types f respnse may ccur. Direct respnse: binding t the receptr directly causes the cellular respnse t the messenger. Secnd messenger invlvement: Binding t the receptr mdifies it, leading t prductin f a secnd messenger, a mlecule that causes the effect. In each case messenger binding induces a cnfrmatinal change in the receptr prtein. Binding f the messenger resembles binding f a substrate t an enzyme in that there is a dissciatin cnstant inhibitin (by antagnists) which may be cmpetitive, nncmpetitive, etc. 2. A variety f messengers can bind t varius tissues. Varius cellular respnses may ccur, depending n the tissue. Either psitive r negative respnses may ccur, even in the same tissue, depending n the type f receptr. 3. The respnse f a cell t a messenger depends n the number f receptrs ccupied. A typical cell may have abut 1000 receptrs. Only a small fractin (10%)f the receptrs need t be ccupied t get a large (50%) respnse. Receptrs may have a dissciatin cnstant f abut 10 exp -11; this is the cncentratin f messenger at which they are 50% saturated. Thus very lw cncentratins f messengers may give a large respnse. The acetylchline receptr f nervus tissue exemplifies a direct respnse type f receptr. 1. The receptr is a cmplex pentameric prtein which frms a channel thrugh the membrane. 2. Mechanism f actin. Binding f acetylchline, a small mlecule, at the exterir surface causes the channel t pen. (Binding) Na + and K + flw thrugh the channel, deplarizing the membrane. (Respnse) The esterase activity f the receptr then hydrlyses the acetylchline, releasing acetate and chline, and terminating the effect. (Recvery) The prcess can nw be repeated. Sme receptrs invlve secnd messengers. Smetimes the binding f an effectr t a receptr leads t the frmatin f an intracellular mlecule which mediates the respnse f the effectr. 1. Definitin: This intracellular mediatr is called a secnd messenger. 2. Effect f secnd messenger frmatin: Since a receptr usually frms many mlecules f secnd messenger after being stimulated by ne mlecule f the riginal effectr, secnd messenger frmatin is a means f amplifying the riginal signal. 3. The frmatin and remval f the secnd messenger can be cntrlled and mdulated. Cyclic AMP (camp) is a secnd messenger that mediates many cellular respnses. 1. Structure f camp: an internal (cyclic) 3', 5'-phsphdiester f adenylic acid. 2. The mechanism f actin f camp is t activate an inactive prtein kinase.
Animated activatin sequence. Since an active prtein kinase which acts n many mlecules f its substrate is prduced, this prcess is an amplificatin f the riginal signal. Since the prtein kinase is activated by camp it is called prtein kinase A. 3. camp is synthesized by the enzyme, adenyl cyclase. The reactin ATP < -> camp + PP i is reversible, but subsequent hydrlysis f the PP i PP i + H 2 O -> 2 P i draws the reactin frward and prevents reversal. Adenyl cyclase is an enzyme, and therefre it is als part f the amplificatin system. camp is degraded by camp phsphdiesterase. camp + H 2 O -> AMP 4. Adenyl cyclase is cntrlled by tw membrane prtein cmplexes, G s and G i. G-prteins are a class f prteins that are s named because they can react with GTP. There are G-prteins in additin t the nes under cnsideratin here. G s and G i are s named because they stimulate and inhibit, respectively, adenyl cyclase. 5. The actin f the G-prteins. Structure: G-prteins are cmplexes f three different subunits, alpha, beta and gamma. Beta and gamma are similar in the G s and G i prteins. The alpha-subunits are different, and are called alpha s and alpha i, respectively. Mechanism: Receptr-messenger interactin stimulates binding f GTP t the alpha-subunits. The alpha-subunit with its bund GTP then dissciates frm the beta-gamma cmplex. The alpha-subunit with its bund GTP then acts n adenyl cyclase. alpha s -GTP stimulates adenyl cyclase. alpha i -GTP inhibits adenyl cyclase. 6. Terminatin f the signal ccurs at several levels. The alpha-subunit f the G-prtein has GTPase activity. After it cleaves the GTP it reassciates with the beta-gamma cmplex t frm the riginal trimer. camp already frmed is cleaved by camp phsphdiesterase. The hrmne gradually and spntaneusly dissciates frm the receptr. Insitl triphsphate (IP 3 ) and diacylglycerl (DG) are als secnd messengers. 1. Animated activatin sequence. 2. IP 3 and DG are synthesized by the enzyme, phsphlipase C, which has phsphatidylinsitl 4,5-bisphsphate (PIP 2 ) phsphdiesterase activity. PIP 2 is a nrmal minr cmpnent f the inner surface f the plasma membrane. 3. The phsphdiesterase is cntrlled by a G-prtein in the membrane, which activates the phsphdiesterase. 4. Mechanism: IP 3 and DG have separate effects. IP 3 releases Ca ++ frm the endplasmic reticulum. The Ca ++ then activates certain intracellular prtein kinases. DG activates prtein kinase c, a specific prtein f the plasma membrane. Nte that bth IP 3 and DG activate prtein kinases, which in turn phsphrylate and affect the activities f ther prteins. 5. Terminatin f the signal ccurs at several levels. IP 3 is hydrlyzed. Ca ++ is returned t the endplasmic reticulum r pumped ut f the cell.
The GTPase activity f the G-prtein hydrlyses the GTP, terminating the activity f the phsphlipase C. 6. Many systems respnd t changes n IP 3 and DG. Be aware f the large number f systems affected. Insulin and grwth factr receptrs. The insulin receptr exemplifies receptrs fr which n secnd messenger has yet been identified. Structure: The insulin receptr is a tetramer with tw kinds f subunits, alpha and beta. Disulfide bridges bind them tgether. The mechanism f signal transmissin is unclear. Many f the cellular respnses are well knwn, e.g. 1. Glucse transprt 2. Prtein phsphrylatin -- Insulin and many grwth factrs activate a prtein kinase which phsphrylates a tyrsyl residue in the target prteins, including the receptr itself. The phsphrylatin f tyrsyl residues is unusual; usually seryl r threnyl residues becme phsphrylated. The significance f this type f phsphrylatin is unknwn. Terminatin f the insulin and certain grwth factr signals invlves internalizatin and degradatin f the hrmne within the cell. 1. The receptr-insulin cmplex migrates t a regin f the plasma membrane with the prtein clathrin cating its inner surface. 2. This regin frms a "cated pit," a regin that invaginates and pinches ff, frming an intracellular "cated vesicle." 3. The cated vesicle fuses with a lyssme; the lyssmal prteases degrade the hrmne specifically, leaving the clathrin and the receptr unharmed. 4. The receptr and clathrin recycle, and are returned t the plasma membrane. Return t the NetBichem Welcme page. jb Last mdified 1/5/95