BIOMG1350 Prelim 1 - L7 Slides - PDF

Prelim 1: Next Monday, Sep 23th All exams are taken on CANVAS in class (12:20 - 1:10 pm): CANVAS/EXAMS/Exam Instruction Check your seat assignment (by Friday Sep 20th): CANVAS/EXAMS – either in Call Auditorium or Klarman G70. Full instructions on Can...

Prelim 1: Next Monday, Sep 23th All exams are taken on CANVAS in class (12:20 - 1:10 pm): CANVAS/EXAMS/Exam Instruction Check your seat assignment (by Friday Sep 20th): CANVAS/EXAMS – either in Call Auditorium or Klarman G70. Full instructions on Canvas/Exams/Exam Instructions. It is your responsibility to read and follow the instructors. Not following the instructions can be considered cheating and may result in 0 points in the Prelim and further penalties. So please don’t!!! Material: lectures 2-7 and sections 2-4 Note: we grade based on material taught in class (lectures and sections) Practice Prelim 1 is available on: CANVAS/EXAMS BIOMG1350 TAs review sessions! Saturday/Sunday 1-4 PM Biotech Racker Room G01 (Instructions in CANVAS/EXAMS) BIOMG1035 review session: Sunday 6-7:30 pm Stimson Hall G01 Office Hours with Martin Graef: Wed 1:30 – 2:30 pm Biotech 201  The Cytoskeleton and Molecular Motors Learning Objectives: Understand the components of the cytoskeleton and their overall organization Understand how microtubules are organized and how molecular motors use ATP hydrolysis to do mechanical work moving organelles along them Understand the structure of actin filaments and how myosins can use them to do mechanical work, like in muscle Watch how this molecular motor can use ATP hydrolysis to walk! Today’s topics: 1.Quick cytoskeleton overview 2.Microtubules and their motors 3.Microfilaments and their motors The cytoskeleton consists of three filament systems that provide shape and structure to cells Cell shape! i.e. keratin Today’s topics: 1.Quick cytoskeleton overview 2.Microtubules and their motors 3.Microfilaments and their motors  Microtubules Centrosome serves as a “MTOC” (microtubule organizing center) icrotubules are hollow tubes of a- and b-tubuli -tubulin -tubulin 13 protofilaments GTP b a b a- and b- tubulin dimers a b a b a b a- and b- tubulin dimers a b a Microtubules have a ‘plus end’ that is favored for assembly, and a ‘minus end’ Figure 17-9 Essential Cell Biology favored for disassembly  Microtubules usually grow out of an MTOC “MTOC” (microtubule organizing center) + + + + + + + + + + + + Centrosome g-tubulin complexes nucleate microtubules and stabilize the minus ends Figure 17-8a Essential Cell Biology  Tubulins are GTP-binding proteins GTP in b-tubulin is slowly hydrolyzed to GDP once incorporated into microtubules bound tubulin is more stably associated with microtubules than is GDP-bound tub allows microtubules to dynamically grow and shrink ubulin bound to GTP: orange ubulin bound to GDP: dark green ubulin bound to GTP: light green Microtubule plus ends can be stabilized in cells by binding to certain proteins Figure 17-16 Essential Cell Biology  Microtubule dynamics are critical for cell division Taxol is a small molecule that binds to and stabilizes microtubules Among other effects, this blocks cell division (more on mitosis later) Taxol is a common chemotherapy drug used to treat breast cancer rotubules transport cargo along a nerve cell a Outward transport occurs up to speeds of about 5µm/s Figure 17-14 Essential Cell Biology  Cytoplasm squeezed from squid giant axon reveals the motion of organelles (name for nerve cytoplasm) Figure 17-19 Essential Cell Biology Organelle movement along MTs in cell extracts Experimental observations: 1. If ATP was removed, the organelles (“cargo”) did not move 2. If ATP was replaced with non-hydrolyzable ATP analog, still did not move These observations tell us: The motors use the energy of ATP hydrolysis (ATP -> ADP + Pi) to move cargos! Movie 17-6 Essential Cell Biology  Motor proteins move along microtubules using their globular heads Cargo Cargo Figure 17-16a Essential Cell Biology fferent motor proteins transport different carg Figure 17-17 Essential Cell Biology tubule motors position organelles in eukaryoti Kinesin motors move ER tubules towards the periphery (edges) of the ce Dynein motors move Golgi membranes towards the center of the cell + + + ER Golgi + + + + MTs MTs + + ++ MTs in green ER in blue MTs: microtubules Figure 17-18 Essential Cell Biology  The structure of kinesin Coiled-coil dimerization region Two ATP-binding Cargo-binding Motor Domain ‘Tail’ ‘Heads' Neck linker two a-helices wrapping around each other Figure 16-62 Molecular Biology of the Cell  Mechanism of kinesin movement Movie 17-7 Essential Cell Biology  Kinesin couples ATP hydrolysis to conformational change coiled-coil stalk ATP ADP trailing leading head head Leading head binds to ATP, which causes a conformational change propelling the trailing head forward by 16 nm. The new leading head with bound ADP, finds a binding site on the microtubule The new leading head releases ADP, and, coordinately, the trailing head hydrolyzes ATP to ADP + Pi  Kinesin couples ATP hydrolysis to conformational change coiled-coil stalk ATP ADP trailing leading Kinesin hydrolyzes one ATP for head head each step along a microtubule ATP vs ADP binding results in two key differences: 1.affinity of head for MT: if ADP is bound, affinity is low if ATP is bound, affinity is high (if no nucleotide, affinity is also high) 2. Position of neck linker relative to motor head is different (conformational change) Kinesin is a highly processive motor coiled-coil stalk ATP ADP trailing leading head head ‘Processive’ means that kinesin can take many steps without dissociating from the microtubule, because one head is always firmly bound as it steps down a microtubule. => kinesin can transport cargo over long distances. Coordination between heads allows processive movement Today’s topics: 1.Quick cytoskeleton overview 2.Microtubules and their motors 3.Microfilaments and their motors Actin filaments in interphase cells ctin filaments are thin, flexible protein thread plus (+) end minus (-) end in filaments grow at the “plus” end of the filam The ATP that is bound to actin is slowly hydrolyzed to ADP Actin monomers dissociate from the minus end In cells, actin polymerization is controlled by actin nucleator proteins Actin-Binding Proteins control the behavior of actin filaments Myosin Skeletal Muscle Muscle cells are pretty big! “Multinucleate” means more than one nucleus per cell. During development, many muscle progenitor cells fuse together to become very big! Sarcomeres are the contractile units of muscle Figure 17-41 Essential Cell Biology  Myosin molecules associate to form bipolar myosin filaments ATPase Head Domains All actin dependent motors belong to the myosin family coiled-coil tail There are many different myosin genes: At least 14 different sub-families of myosin genes have been identified! Muscle myosin belongs to the myosin-II subfamily Figure 17-38 Essential Cell Biology Each Myosin head walks along actin filaments due to a cycle of ATP binding and hydrolysis ATP binding causes the release of myosin from actin fila ATP hydrolysis causes conformation change, and the myosin head with ADP and Pi tightly bound moves into a new position (“cocked”) Figure 17-43 Essential Cell Biology Each Myosin head walks along actin filaments due to a cycle of ATP binding and hydrolysis Weak binding of the myosin head to a new site on the actin filament causes the release of phosphate Phosphate release results in another conformation ch called the “power stroke”, which generates a force and moves the actin filament relative to the myosin filament ADP is released and myosin remains attached to the actin filament, ready for another cycle. We say this is not processive because each motor head works independently Figure 17-43 Essential Cell Biology  Muscles contract by a sliding filament mechanism ++ - - ++ ++ - - ++ ++ - - ++ ++ - - ++ Figure 17-42 Essential Cell Biology