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BCOR 103:Cytoplasm

Cytoskeleton function
1. scaffolding 2. organelle positioning 3. support for organizing machinery of macromolecular metabolism 4. movement of materials and organelles 5. locomotion 5. transduction of extracellular signals
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The "cell skeleton" is made up of __.
- architectural nucleus proteins
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The cytoskeleton is made up of: (2)
1. cytoplasmic filaments and their associated proteins/factors 2. material that remains between the nuclear envelope and plasma membrane after cell lysing and washing
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primary set of cytoskeleton filaments (4)
1. microtubules 2. intermediate filaments 3. microfilaments 4. thick filaments
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microtubules
-hollow tubulin tubes -used in division and intracellular support
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intermediate filaments are __ that ___.
- rope like fibers - resist mechanical stress
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Microfilaments are made up of __ and involved in ___.
-globular chains of actin protein -involved in cellular elasticity and motility
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thick filaments are made up of __ and are used in ___.
-rough, bipolar aggregates of myosin protein -contractile activities of cells
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Isoforms 
-different distinct functional versions of the cytoskeleton -easily broken down and rebuilt
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general areas of eukaryotic cytoplasm (2)
1. cortical cytoplasm 2. subcortical cytoplasm/endoplasm
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Cortical cytoplasm structure/location
- Mostly microfilaments -thin, gel like area beneath the plasma membrane
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cortical cytoplasm function
- exclude other organelles from the region - maintain cell shape
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Subcortical cytoplasm/endoplasm
-syrup like -most organelles found here
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nuclear lamins
-subgroup of intermediate filaments present in all eukaryotic nuclei
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Cyotplasmic IFs are found in __ but not in __.
-mammalian animal cells - plant cells
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Subtypes of IFs (5)
1&2. keratins 3. Desmin, vimentin, GFAP 4. (neurons) 3 neurofilament proteins combined as a homo/heterodimer 5. copolymers of nuclear lamins A,B,C
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IF protein structure
-N and C terminal globular domains -central rod domain
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Essentially all of the differences in the 5 IF subtypes exist in the ____.
globular domains
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Globular domain function
- controls the assembly of proteins - connects intermediate filaments with non-IFs
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central rod structure
-alpha helical heptad repeat sequence -six steps in assembly
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6 steps of central rod structure assembly
1. formation of coiled-coil dimer 2. anti-parallel and half staggered dimer association into a tetramer 3. end to end association of tetramers into "protofilament" 4. pairs of protofilaments associate laterally into an "eight chain"protofibril 5. four protofilaments wrap around each other, forming "cylindrical 10nm IF filament" 6. IFs may bundle into "tonofibrils"(c,t,p,e,c,t)
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tonofibrils
- hemidesmosomes attach to intermediate filaments of cytosketon
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IF network regulation
-"pool"of ready dimers/tetramers to make IFs -Assembled/disassembled quickly in cycles through phosphorylation/dephosphorylation
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IF function
-increase efficency and provide protection -greatest resistance to stress -position organelles -organize sarcomeres -support membrane
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microtubules are comprised of ___ subunits.
tubulin
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Tubulin is made of ___ and ____ ____. 
Alpha beta subunits
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Tubulin subunits are __ ___ proteins.
guanine binding
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When alpha and beta tubulin subunits associate, they form a 
protofilament
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___ are formed when __ tubulin protofilaments bind to form a hollow cylinder
Microtubule 13
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Microtubule associated proteins (MAPs)  
- specifically bound materials on the surface of microtubules/alpha beta subunits
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MAP subgroups (3)
1. Structural 2. Specialized 3. Microtubule based motors
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Structural MAP function
Enhance properties of their microtubule
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Specialized MAP function
Proteins that are unique to a distinct MT function of a specialized cell type
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Microtubule based motor function
Use microtubules as "tracks" for cellular motility
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3 stages of MT formation
1. Lag 2. Elongation 3. Steady state
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Lag phase
- dependent on MAP proteins - ring of tubulins form
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Elongation phase
addition of the alpha/beta dimers to the ends of the protofilaments elongating the MT
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steady state phase 
-Length of microtubule (MT) is in "equilibrium" with the amount of unpolimerized alpha/beta subunits
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Critical concentration
The concentration of free tubulin (a/b subunits) at the steady state of MT formation
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If the free tube concentration goes down, the MT length __
goes down
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Microtubule organizing center (MTOCs)
distinct areas where MTs originate
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Important properties of MTOCs
1. synthesize new MTs, even when free tubulin is below critical concentration 2. hold the slower growing (-) end of MTs, allowing more growth at the faster end (+) 3. MTOC synthesis is saturable and regulatable
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MTOC types/examples (3)
1. Basal bodies (cilia/flagella anchor) 2. spindle pore body (organize mitotic spindle in yeast) 3. centrosome
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centrosome structure
-Pair of centrioles -pericentriolar material
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MTs originate from ___ in centrosomes
-pericentriolar material 
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What is dynamic instability
- polymerized and free tubulin are in balance - some MTs grow while others shrink
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What do MTOCs do during interphase
send out MTs in all directions
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MTs can grow outside of the ___ ___.
cell membrane
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If MTs remain stable long enough, they: 
are chemically modified with MAPs
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When do MTOCs/centrosome duplicate?
at the beginning of mitosis
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Aster MTs
- MT generated in a cell with to MTOCs - shorter
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Chromsomes act as a __ on MTs
Cap
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Microtubule motors
- couple vesicles to an MT and move them faster than diffusion
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Classes of microtubule motors (2)
1. kinesin class 2. dynein class
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Kinesin class motors
- Two heavy chains that house two globular ATPase motor heads - Two light chains that form the cargo-carrying tail. - "walk"down the MTs with the cargo
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Dynein class motors
- Active proteins in the movement of cilia and flagella - this class moves toward the - end of MTs
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Primary protein subunit of microfilaments is
Actin
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Describe actin structure
- Globular protein - Bi-lobed protein which binds ATP in the cleft
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- Globular protein - Bi-lobed protein which binds ATP in the cleft
- double helix - polymerized actin monomers
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microfilaments are __ in diameter
6 nm
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T/F microfilaments are the longest structural subunits
False; shortest
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similarities between MTs and microfilaments
- both have a + growing end and a - slow growing end
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Describe treadmilling 
-found in microtubules - subunits are taken from - end and added to + end
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microfilaments alone don't strongly associate with anything. In order to perform their function they need __ and __ __ __.
- microfilament - actin associated proteins
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Families of microfilament associated proteins (4)
1. monopolar binders 2. capping proteins 3. linking proteins 4. motor proteins
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Monomer binder function
- only bind to free actin - regulates microfilament formation
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3 jobs of capping proteins
1. cap one end of microfilament (usually + end, confining growth to - end) 2. sever the microfilament by binding to the side of it 3. nucleation creates a new filament growth protein
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Linking protein subclasses (3)
1. bundlers hold microfilaments side by side for additional strength (myosin, microvilli) 2. cross linkers put microfilaments across one another, forming a gel 3. membrane linkers connect microfilaments to the membrane by their end or side
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Motor proteins are also known as:
myosins
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Two groups of motor proteins
1. Myosin I 2. Myosin II
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Describe Myosin II
-Myosin in muscle - Made of 2 heavy chains, 2 light chains, and 2 regulatory light chains - 2 globular heads and a tail - view figures for how chains move
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Describe Myosin I
- 1 motor head - no tail - end of these molecules has binding sites for either a filament or membrane lipids
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Myosin I function
Myosin I function
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4 stages of locomotion
1. polarity determination 2. leading edge protrusion 3. substrate adhesion 4. traction/cell body positional change
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Describe polarity determination
- occurs in response to an orientated signal - dependent upon the capture and orientating of the microtubule network
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Describe protrusion
Microspikes that probe foreward
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Microspikes
a thin tube of cytoplasm supported by a bundle of actin filaments or lamellipodia
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lamellipodia
a broad thin band of cortical cytoplasm packed with a meshwork of actin filaments
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3 proposed mechanisms for the generation of the protrusive force that is required to extend the protrusions forward?
1. Actin polymerization: rapid growth of filaments pushes membrane outward 2. actin/myosin/membrane interaction: myosin I propels actin rods against membrane 3. Gel osmotic forces: concentration of particles below membrane brings in water and expands membrane
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Describe substrate adhesion
when protusions encounter an adhesive region of the substratum it will adhere, stabilizing foreward expansion
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Describe traction/body movement that shifts cell towards protrusion
1. stress fiber contraction (rear of the cells contract due to sarcomere-like stress fibers, pulling cell foreward) 2. Motor protein induced cytoplasmic flow (fluid is "rowed" forward by minimyosins) 3. general cortical tension (cytoplasm pushed foreward)
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IF subtypes I and II and both __ which exist as __ and are only expressed in __ _.
- keratins - heterodimers - epitheilal cells
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Mitochondria can _ _ causing elongated organelle networks
fuse together
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4 structures of mitochondria
1. Outer membrane 2. intermembrane space 3. inner membrane 4. the mitochondrial matrix
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1. Outer membrane 2. intermembrane space 3. inner membrane 4. the mitochondrial matrix
-high concentration of mitochondrial porin (form beta barrels) - 5000MW substances or smaller can pass through
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intermembrane space
-between outer and inner membranes - metabolic content roughly matches cytosol, but with different proteins (and fewer)
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primary protein found in intermembrane space
-electron transporter cytochrome-c
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general description of inner membrane
-Folded into cristae to increase surface area, with more cristae for greater energy needs. - Not smooth, has many projections. -Very impermeable. -High protein content (transporters, electron carriers, enzymes)
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Two regions of inner membrane
1. inner boundary membrane 2. cristae proper
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inner boundary membrane 
directly opposes the outer membrane, rich in metabolite transporters and protein translocators 
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Cristae proper
Rich in electron transport chain compoents and ATP-synthase
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cristae junctions
specialized membrane constrictions that attach the cristae proper to the inner boundary membrane
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mitochodrial matrix
- fluid component of the mitochondria -densely packed with proteins and other particles -Contains enzymes and metabolites of the citric acid cycle and beta-oxidation. -Also contains the DNA and ribosomes that make the mitochondria semi-autonomous.
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The electron transport chain uses __ from __ __ of __ to produce __ which is subsequently used to produce __.
-electrons -oxidative reactions -catabolism - electrochemical, energy storing gradient -ATP
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What does the electron transport chain do with the electrons
What does the electron transport chain do with the electrons
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steps using complex 1 of the electron transport chain
- over 20 subunit proteins embedded in inner membrane -Receives an electron from NADH, creating NAD- and releasing H+ into the matrix. Electrons pass one-at-a-time to a molecule of Coenzyme Q (CoQ). Movement of electrons releases energy that allows the complex to pump 4 H+ from the matrix to the intermembrane space.
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steps using complex 2 of the electron transport chain
Succinate is oxidized and its electrons/hydrogens are transfered to FAD producing FADH2. Complex 2 releases H+ into the matrix, and transferers electrons to CoQ, but lacks the energy to transport H+ across the membrane.
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Steps using complex 3 of the electron transport chain
Complex 3 has a large binding pocket for CoQH2, which binds and releases its H+ to the intermembrane space. The complex then transfers one electron to Cytochrome-c and one to a different CoQ molecule. This happens twice, providing a second electron to the second CoQ molecule and allowing it to carry two H+ to the intermembrane side
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What is the name of the cycle that complex 3 is involved in?
Q-cycle
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The Q cycle allows CoQ to shuttle how many hydrogens to the intermembrane space per pair of electrons transported along the chain? 4 H+ per electron
4 H+ per electron
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Steps using Complex 4 of the electron transport chain
The electrons from cytochrome-c (a mobile electron carrier) are transported from complex 3 to 4. Subunit is used for binding molecular oxygen, transferring electrons to the oxygen creating reactive oxygen species, absorbing H+ from the matrix and uniting them with reactive oxygen to form water, and using the energy released in the process to pump 2H+ from the matrix to the intermembrane space for every pair of electrons transported.
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primary function of the electron transport chain is to 
Move H+ from the matrix to the intermembrane space
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When an electron pair from NADH is inserted into electron transport chain, how many H+ are transferred from which complexes
-4 are transported by complex 1 -4 are transported by the Q-cycle (complex 3) -2 are transported by complex 4 - 10 in total
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when an electron pair is transferred from FADH2, how many H+ are transported from which complexes?
- 6 H+ are isolated from complex 2, which are not transferred to the intermembrane space until complex 4
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Proton-motive force
- The name for dual concentration and electrical gradient across the inner membrane - utilized to create ATP
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FoF1 synthase is a __ type pump used in to produce _ using the __ __.
- F - inner membrane - ATP - H+ gradient
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3 steps for the production of ATP using FoF1 ATP synthase
1. H+ is transferred from the intermembrane space to the matrix through the pore in synthase. This causes the c-ring of the protein to rotate. 2. As this part rotates, it causes a rotor unit to rotate as well. The rotation of the rotor causes a conformational change in the beta subunits of the synthase 3. The confrontational change of the beta subunit form the ATP and release it.
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How many ATP molecules can be produced from one glucose molecule?
How many ATP molecules can be produced from one glucose molecule?
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How many ATP molecules are released per rotation of the rotor in FoF1 ATP synthase?
How many ATP molecules are released per rotation of the rotor in FoF1 ATP synthase?
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What are the 3 conformations of the beta subunits in FoF1 ATP synthase
1. Loose (allows the binding of ATP and Pi 2. Tight (combines two previous substrates into ATP) 3. Open (release ATP)
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