bibc 100 handout 4
DESCRIPTION
I've included material you will be covering Tues 4/26 which will be everything up to the Midterm. I just realized section will be at the same time as Review Session on Monday so feel free to go or stay in section. If you do come, we will work on the Handout and go over some Midterm practice problems. If no one comes after 15min, I will be studying in CLICS till 1am x_x. Feel free to stop by if you have questions!Hope studying is going well!TRANSCRIPT
BIBC 100 – Structural Biochemistry TA: Lydiesther MartinezDr. Montal [email protected] ’11 Section: Mon 7pm
WEEK 4 and 5Protein Folding Introduction
o Problem: protein folding is statistically impossible if it is completely ____________o Levinthal’s paradox – proteins usually fold in 10-1 to 103 seconds
o Conclude that folding is not ____________, but deterministico USA – ____________ (action), ____________ (energy), ____________ (kinetics)
o Important because folding is 2nd genetic code – could lead to computer designed proteins
Anfinsen Experiment Tested ribonuclease to see if protein folding was dictated by the ____________sequence Reagents:
o ____________– interact with water and other molecules to disrupt hydrophobic interactionso -____________– reducing agent to break disulfide bonds
Found that primary sequence was enough for small globular proteins to fold into ____________state
Free Energy Funnel for Protein Foldingo Entropy – drives towards unfolded state with hydrophobic residues
exposedo ____________– drives toward folded state with hydrophobic
interactions, H bonding, ionic interactionso Free energy is the difference – drives towards folded stateo Common obstacles to folding:
____________of hydrophobic regions Non-native ____________bridge formation ____________of proline
o Unfolded molten globule (loose 3° structure) is fasto Molten globule native structure is slowero Some proteins never reach their native stateo Other stable structures are not functional
transition state between molten globule and native state requires ____________in free energy
Prions & Alzheimer’s Diseaseo Prion – “proteinaceous infectious only” o PrPc (prion protein) is a normal constituent of cells with significant helix contento Changes in conformation allow____________ content (PrPsc) propagates to
other proteins forms amyloid fibers. Fibers accumulate in nerve cells, which spend all their energy trying to get rid of the toxin lose homeostatic energy
o Results in loss of memories death
Protein Folding Chaperones – GroEL/GroES Complex ____________ (HSP) – prevent damage to proteins in response to high levels of
heat by preventing aggregation of exposed hydrophobic regionso Nonspecific – recognizes exposed hydrophobic patches
HSP 70: regulates translocation to the ER, assembly & disassembly of oligomers GroEL (Hsp 60): 7 subunits with apical (______), intermediate (___), equatorial domains (____) GroES (Hsp 10): ___ hairpin roof – sheet – mobile _______(integrated with GroEL) GroEL/ES complex has 2 barrels end-to-end, so each can accommodate a protein but GroES cap is an
BIBC 100 – Structural Biochemistry TA: Lydiesther MartinezDr. Montal [email protected] ’11 Section: Mon 7pm
__________ inhibitor for the other barrel
Protein Stability o Can detect through _________ (site-directed protein engineering)o Disulfide (S-S) bridges: reducing the entropy through covalent bonds increases protein stabilityo G & P: Glycine is much more flexible than Proline (side chain is covalently bonded to main chain)
o G P or A increases stabilityo Dipole stability: N terminus should have (-); C terminus should have (+) to increase stabilityo Hydrophobicity in core cavity increases stability
(thermodynamics)
Allosteric Control o Allosteric effectors: small ligands that bind to
_________ _________ recognition sites (away from functional binding sites)
o Trp repressor controls operon that synthesizes L-trpo Absence of trp: helices tilt inward - 29Å distance
between binding sites (inactive repressor)o trp bound (2) recognition helices ______ apart for proper binding of DNA (active repressor)
CAP – catabolite gene activating protein Motif: _________ _________ DNA binding protein that assists RNA polymerase in effective binding
o Promotes initiation of RNA synthesis Functions as _________ Activated by _________ (allosteric effector) Interacts directly with base pairs & phosphate groups Bending of DNA contributes to activation of transcription _____ – mutations have been associated with many diff types of cancer
(WT is tumor suppressor by controlling cell cycle)o DNA binding protein interaction of helix with groove of DNAo H-bonds between protein & base pairs
Zinc Finger Motif o Motif: 1 _________ & 2 _________ coordinated by a ____ metal
(stabilized by 2 Cys, 2 His – midterm 6a) Approximately 30 amino acids Highly conserved coordination sites (midterm 6b)
BIBC 100 – Structural Biochemistry TA: Lydiesther MartinezDr. Montal [email protected] ’11 Section: Mon 7pm
Proteins contain anywhere from 1-60 zinc finger motifs within sequence alpha helix side chains _________ bond with base pairs of DNA (midterm 6c) Fig 10.3 – schematic diagram of 3 Zn fingers binding in tandem to major groove – surround DNA
Leucine Zipper Motif Motif: _________ _________ alpha helices Heptad repeat: almost all 4th residue is _________ (fig 10.17)
o Character of AA repeat every ___ AA _________ core is major contributing factor to stability Distal, basic region on each helix interacts with major groove
o Increased concentration of _________o Likely AA: _________ and _________
Immune Response Immune system: recognizes, destroys, and remembers _________ _________
while ignoring _________ cells & proteins Antigen recognizes foreign molecules immune response
o ____ cells release immunoglobulins to circulationo ____ cells have cell receptor that recognizes _________ protein
Immunoglobulin Fold Heterotetramer: 2 ____ chains + 2 _____ chains
o Linked by ____ disulfide bonds stabilityo Hinge region of HC _________ (Y/T conformations)
Ig fold is distorted _________o Each LC contains _____ Ig folds, each HC contains ___ Ig foldso Distorted conformation stabilized by _____ disulfide bridges
Antigen recognition at loops between strands at N-terminus of Ig Each Ig recognizes ___ antigen(s)
o Different B cells diff Igo Heavy & light chains are encoded by different geneso N terminus regions contain the most variabilityo Hypervariable loops provide antigen recognition & specificity
Bind to antigen via _________ bonds Papain cleaves _________ bonds and separates the IgG into 2 F__ sites (light & heavy chain – antigen
binding site) and F___ (2 heavy chains)
Good luck!