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Cell Biology Course Pack.docx

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Department
Biology
Course Code
Biology 2382B
Professor
timeshenko

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Cell Biology – Course Pack Microtubules – 25 nm, Microfilaments (Actin!) – 7-9 nm, Intermediate Filaments – 10 nm Microtubules - Polymer of α- β- monomers (55 kDa each). Found in dimer (110 kDa each and 8nm long!) because it is more stable. - Dimers form protofilaments which then form hollow tubes. 25 nm diameter and up to 100 μ m long. - α- is (-) end and β- is (+) = polarity! Grow preferentially from (+) end. - Both α- β- monomers can bind GTP, however ONLY β-monomer can hydrolyze the GTP into GDP. GTP creates stronger bonds, once hydrolyzed, easier to disassemble - Protofilaments can arrange into singlets (13 protofilaments that are relatively unstable, constantly rearranged) or doublets (13+10, found in cilia and flagella – axonemel microtubules…nothing to do with axons) or triplets (13+10+10, found in basal bodies and centrioles). Centrosome: Is the Microtubule Organizing Center (MTOC) in mammals. Centrosomes are not found in plants. Made of centrioles which are nine triplets at 90° to each other. Note that microtubules do not arise directly from centrioles but rather from γ-tubulin (specifically, γ-TuRC, γ-tubulin ring complex provides nucleating sites) and the protein pericentrin. - Another type of MTOC are basal bodies at the base of flagella and cilia. - There is a critical concentration (Cc) of dimers above which you get polymerization and below which you get depolymerization - Also influenced by temperature - Treadmilling may occur if Cc is reached at one end but not the other. However, this is done more so with actin, fairly rare with microtubules. Also note that Cc may be different for respective ends (e.g. (-) end has higher Cc, however, it is usually capped and does not usually undergo growing and shrinking) - Important, microtubules are dynamic! Constant remodeling in the cell. Microtubule Disrupting drugs: colchicine depolymerizes and taxol which stabilizes. Both are bad for the cell (thus why they’re often used as anti-cancer drugs, disrupt cellular replication and other processes), remember dynamics (growing AND shrinking) are very important! Microtubule Associated Proteins: Stabilize microtubules. Map2 and Tau. Eb1 is a MAP which also acts as an attachment site Microtubule Binding and Severing Protein (1)Kinesin-13 – a protein that requires energy to disassemble (uses ATP) (2)Stathmin – does not require energy, binds, and promotes GTP hydrolysis of β- subunit. (3)Katanin (sword) – enzyme (so it requires ATP) Microtubules as Tracks for Transport – How do we know? Experiments using Squid axon, it’s large and very long. Inject radiolabeled amino acids into squid axon. The amino acid will be incorporated in protein. This will allow us to track movement at various time points. Kinesin  most are (+) end directed (anterograde) motor proteins. Made of 2 heavy chains containing ATPase activity (associate with the microtubules containing head, neck and stalk) and 2 light chains which are variable in order to bind variable cargo. Type 1 – Conventional. Moves 16 nm each hydrolyzed ATP (remember, each dimer is 8 nm long) Type 2 – Heterotrimeric Type 5 – Homodimeric – does not carry cargo! Thus, no light chains. Used for filament sliding. Type 13 – Has head domain, neck domain, but no stalk domain. Cannot bind cargo, used for end disassembly as previously mentioned. Dynein (-) directed (retrograde) motor proteins. Dynactin Binding Domain binds the dynactin heterocomplex whic
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