Bài giảng Biochemistry 2/e - Chapter 17: Molecular Motors

Chapter 17Molecular Motorsto accompanyBiochemistry, 2/ebyReginald Garrett and Charles GrishamAll rights reserved. Requests for permission to make copies of any part of the work should be mailed to: Permissions Department, Harcourt Brace & Company, 6277 Sea Harbor Drive, Orlando, Florida 32887-6777 Outline17.1 Molecular Motors17.2 Microtubules and Their Motors17.3 Skeletal Muscle Myosin and Muscle Contraction17.4 A Proton Gradient Drives the Rotation of Baterial FlagellaTubulin and MicrotubulesFundamental components of the eukaryotic cytoskeleton Microtubules are hollow, cylindrical polymers made from tubulin dimers 13 tubulin monomers per turn Dimers add to the "plus" end and dissociate from the "minus" end as in Figure 17.3 Microtubules are the basic components of the cytoskeleton and of cilia and flagella Cilia wave; flagella rotate - ATP drives both! Microtubules in Cilia & FlagellaMTs are the fundamental structural unit in cilia and flagella (see axoneme structure, Fig 17.5) Dynein proteins walk or slide along MTs to cause bending of one MT relative to another Dynein movement is ATP-driven See Figures 17.6 and 17.7 MicrotubulesHighways for "molecular motors" MTs also mediate motion of organelles and vesicles through the cell In axons, dyneins move organelles + to -, i.e., toward the nucleus Kinesins move organelles - to + , i.e., away from the nucleus See Figure 17.8 and compare (a) and (b)Polymerization InhibitorsTherapeutic agents for gout and cancer Colchicine, from autumn crocus, inhibits MT polymerization, mitosis and also white cell movement - it is a remedy for gout and an inducer of larger, healthier plants Vinblastine, vincristine also inhibit MT polymerization - anticancer agents Taxol, from yew tree bark, stimulates polymerization, stabilizes microtubules and inhibits tumor growth, (esp. breast and ovarian)Morphology of MuscleFour types: skeletal, cardiac, smooth and myoepithelial cells A fiber bundle contains hundreds of myofibrils that run the length of the fiber Each myofibril is a linear array of sarcomeres Each sarcomere is capped on ends by a transverse tubule (t-tubule) that is an extension of sarcolemmal membrane Surfaces of sarcomeres are covered by SRWhat are t-tubules and SR for? The morphology is all geared to Ca release and uptake! Nerve impulses reaching the muscle produce an "action potential" that spreads over the sarcolemmal membrane and into the fiber along the t-tubule network What are t-tubules and SR for? The morphology is all geared to Ca release and uptake! The signal is passed across the triad junction and induces release of Ca2+ ions from the SR Ca2+ ions bind to sites on the fibers and induce contraction; relaxation involves pumping the Ca2+ back into the SRMolecular Structure of MuscleBe able to explain the EM in Figure 17.12 in terms of thin and thick filaments Thin filaments are composed of actin polymers F-actin helix is composed of G-actin monomers F-actin helix has a pitch of 72 nm But repeat distance is 36 nm Actin filaments are decorated with tropomyosin heterodimers and troponin complexes Troponin complex consists of: troponin T (TnT), troponin I (TnI), and troponin C (TnC)Structure of Thick FilamentsMyosin - 2 heavy chains, 4 light chains Heavy chains - 230 kD each Light chains - 2 pairs of different 20 kD chains The "heads" of heavy chains have ATPase activity and hydrolysis here drives contraction Light chains are homologous to calmodulin and also to TnC See structure of heads in Figure 17.16Repeating Elements in MyosinThe secret to ultrastructure 7-residue, 28-residue and 196-residue repeats are responsible for the organization of thick filaments Residues 1 and 4 (a and d) of the seven-residue repeat are hydrophobic; residues 2,3 and 6 (b, c and f) are ionic This repeating pattern favors formation of coiled coil of tails. (With 3.6 - NOT 3.5 - residues per turn, a-helices will coil!)More Repeats!28-residue repeat (4 x 7) consists of distinct patterns of alternating side-chain charge (+ vs -), and these regions pack with regions of opposite charge on adjacent myosins to stabilize the filament 196-residue repeat (7 x 28) pattern also contributes to packing and stability of filaments Associated proteins of Muscle-Actinin, a protein that contains several repeat units, forms dimers and contains actin-binding regions, and is analogous in some ways to dystrophin Dystrophin is the protein product of the first gene to be associated with muscular dystrophy - actually Duchennes MD See the box on pages 548-549DystrophinNew Developments! Dystrophin is part of a large complex of glycoproteins that bridges the inner cytoskeleton (actin filaments) and the extracellular matrix (via a protein called laminin) Two subcomplexes: dystroglycan and sarcoglycan Defects in these proteins have now been linked to other forms of muscular dystrophyThe Dystrophin ComplexLinks to disease -Dystroglycan - extracellular, binds to merosin (a component of laminin) - mutation in merosin linked to severe congenital muscular dystrophy -Dystroglycan - transmembrane protein that binds dystrophin inside Sarcoglycan complex - , ,  - all transmembrane - defects linked to limb-girdle MD and autosomal recessive MDThe Sliding Filament ModelMany contributors!Hugh Huxley and Jean Hanson Andrew Huxley and Ralph Niedergerke Albert Szent-Gyorgyi showed that actin and myosin associate (actomyosin complex) Sarcomeres decrease length during contraction (see Figure 17.22) Szent-Gyorgyi also showed that ATP causes the actomyosin complex to dissociateThe Contraction CycleStudy Figure 17.23! Cross-bridge formation is followed by power stroke with ADP and Pi release ATP binding causes dissociation of myosin heads and reorientation of myosin head Details of the conformational change in the myosin heads are coming to light! Evidence now exists for a movement of at least 35 A in the conformation change between the ADP-bound state and ADP-free state Ca2+ Controls ContractionCa2+ Channels and Pumps Release of Ca2+ from the SR triggers contraction Reuptake of Ca2+ into SR relaxes muscle So how is calcium released in response to nerve impulses? Answer has come from studies of antagonist molecules that block Ca2+ channel activityDihydropyridine ReceptorIn t-tubules of heart and skeletal muscle Nifedipine and other DHP-like molecules bind to the "DHP receptor" in t-tubules In heart, DHP receptor is a voltage-gated Ca2+ channel In skeletal muscle, DHP receptor is apparently a voltage-sensing protein and probably undergoes voltage-dependent conformational changes Ryanodine ReceptorThe "foot structure" in terminal cisternae of SR Foot structure is a Ca2+ channel of unusual design Note structure in Figures 17.27 and 17.28 Conformation change or Ca2+ -channel activity of DHP receptor apparently gates the ryanodine receptor, opening and closing Ca2+ channels Many details are yet to be elucidated! Ca 2+ Regulates ContractionTropomyosin and troponins mediate the effects of Ca2+ See Figure 17.29 In absence of Ca2+, TnI binds to actin to keep myosin off TnI and TnT interact with tropomyosin to keep tropomyosin away from the groove between adjacent actins But Ca2+ binding changes all this!Ca 2+ Turns on ContractionBinding of Ca2+ to TnC increases binding of TnC to TnI, simultaneously decreasing the interaction of TnI with actin This allows tropomyosin to slide down into the actin groove, exposing myosin-binding sites on actin and initiating contraction Since troponin complex interacts only with every 7th actin, the conformational changes must be cooperative Binding of Ca 2+ to Troponin CFour sites for Ca2+ on TnC - I, II, III and IV Sites I & II are N-terminal; III and IV on C term Sites III and IV usually have Ca2+ bound Sites I and II are empty in resting state Rise of Ca2+ levels fills sites I and II Conformation change facilitates binding of TnC to TnISmooth Muscle ContractionNo troponin complex in smooth muscle In smooth muscle, Ca2+ activates myosin light chain kinase (MLCK) which phosphorylates LC2, the regulatory light chain of myosin Ca2+ effect is via calmodulin - a cousin of TnC Hormones regulate contraction - epinephrine, a smooth muscle relaxer, activates adenylyl cyclase, making cAMP, which activates protein kinase, which phosphorylates MLCK, inactivating MLCK and relaxing muscleSmooth Muscle EffectorsUseful drugs Epinephrine (as Primatene) is an over-the-counter asthma drug, but it acts on heart as well as on lungs - a possible problem! Albuterol is a more selective smooth muscle relaxer and acts more on lungs than heart Albuterol is used to prevent premature laborOxytocin (pitocin) stimulates contraction of uterine smooth muscle, inducing labor