1 / 58

Protein Function/Enzyme Regulation/Biosignalling

Protein Function/Enzyme Regulation/Biosignalling. Chpts. 5, 6, 12. 1 o , 2 o , 3 o , 4 o Structures REMEMBER??. Structure defines function If >1 polypeptide chain, 4 o structure Chains not independent Book: Proteins are dynamic structures. Definitions (Chpt. 5).

candy
Download Presentation

Protein Function/Enzyme Regulation/Biosignalling

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. Protein Function/Enzyme Regulation/Biosignalling Chpts. 5, 6, 12

  2. 1o, 2o, 3o, 4o Structures REMEMBER?? • Structure defines function • If >1 polypeptide chain, 4o structure • Chains not independent • Book: Proteins are dynamic structures

  3. Definitions (Chpt. 5) • Ligand – bound reversibly to prot • Binding site – where ligand binds • Complementarity • Induced fit – protein flexes  greater complementarity • (What do all of these characteristics remind you of?)

  4. Protein/Ligand Binding • May be regulated by another mol • May be 2nd ligand w/ 2nd binding site • Second ligand interaction  flexing  change in first ligand’s ability to bind • 1st ligand may bind better or worse under influence of 2nd ligand

  5. Hemoglobin (Hb) • 4 polypeptide chains + 4 heme grps • MW 64,5000 old book

  6. Protein (Globin) • Globular • 4 hydrophobic pockets • 4o structure due to interaction ~30 aa’s • a/b subunits interact (not a/a or b/b) • Mostly hydrophobic interactions, some ionic

  7. Heme • Protophorphyrin ring • Binds single Fe as Fe+2 • Sim to pigments • Resonance (electron transport, color, UV absorbance)

  8. Fe Held in Heme; Heme Held in Hb • 6 Coordination sites for heme Fe • 4 bind N’s of protoporphyrin ring • 1 binds globin His R grp N • 1 binds O2 •  Changes in heme electronic prop’s •  Color change • Fe can also bind CO

  9. Globin may be in T state or R state • T state more stable w/out O2 • O2 prefers to bind globin in R state (either poss.) • Bound O2 stabilizes R

  10. O2 Binding to Heme Influences Globin • T state (stable deoxyHb) binds O2 •  Shift in globin conform’n • a, b subunits slide, rotate • b subunits closer • R state results • O2 binding @ Fe  incr’d planarity of heme  altered interactions of R grps of nearby aa’s

  11. Imptc to Hb Function (Transport O2) • O2 must rev’ly bind Hb, but tight enough for transport • Binding of 1 O2 molecule causes TR • R state now stabilized • Subunits have been effected • Now easier for 2nd O2 to bind • 3rd, 4th O2’s • R state strengthened w/ each O2 added

  12. Allosteric Protein • Ligand binding @ one site affects binding abilities @ other sites on same protein • Due to conform’l changes  altered binding site(s) • Hemoglobin example • O2 = activator (stimulator) • Positive cooperativity among subunits

  13. Can be treated mathematically • Similar to Ka, Kd • P + nL  PLn • P = Protein • L = ligand • q= binding sites occ’d/total binding sites • Based on fraction of binding sites occupied, derive Hill coefficient (hH) • = 1, no cooperative binding • < 1, negative cooperativity • > 1, positive cooperativity

  14. Models for Cooperativity • Monod • “All or nothing” • No subunit in any independent conformation • Ligand binds any, but prefers one

  15. Koshland (Sequential) • Subunits more independent • One subunit acts as modulator • Conform’l change influences conform’l changes in other subunits • “Graded effects”

  16. (Sickle Cell Anemia • Mutation  single improper aa in Hb globin • b chain Glu  Val • Now – charge  uncharged side chain •  “sticky” hydrophobic pt @ outer Hb surface • DeoxyHb mol’s associate w/ each other •  Strand, fiber form’n •  Long, thin crescent rbc’s)

  17. Allosteric Effects Regulate Some Enz Activity in Metabolism • Metab pathways mediated by enzymes • Several rxns in succession • Each rxn catalyzed by partic enzyme • P rxn 1 becomes S for rxn 2, etc.

  18. Enzymes Can Be Inhibited • Product inhib’n • Enz may be inhib’d by its own P • Inverse relationship of [P] and further P synthesis • P acts as competitive inhibitor • Resembles S • Fits enz active site • Competes • Inhib’n overcome

  19. Feedback inhibition • Enz may be inhib’d by metabolite from further down pathway • L-ileu prevents form’n • Inhibits thr dehydratase • No other • Thr dehydratase = regulatory enzyme • Regulates pathway

  20. Regulatory Enzymes • Catalyze slowest step • Stim’d or inhibited • Commonly 1st • Point of commitment • May be allosteric OR controlled by covalent modification

  21. Allosteric Regulatory Enzymes • REMEMBER how Hb worked • Modulated • >1 binding site • S binding to one site affects other binding site(s) • Both need not be catalytic • Often 1 regulatory • Both specific for either S or modulator • Often on diff subunits

  22. Modulator binding at regulatory site  conform’l change at catalytic site • May be harder or easier for S to bind • Conform’l changes due to noncovalent interactions

  23. Altered Kinetics of Allosteric Enzymes • M-M model hyperbolic • Allosteric model sigmoidal • If modulator stimulates, more hyperbolic • If modulator inhibits, more sigmoidal • KM changes

  24. Covalently Modified Regulatory Enzymes • Also controlled through modulators • Now modulator covalently bound • At some funct’l grp of aa of enz 1o structure • Need OTHER enz’s to catalyze binding of modulator • Need EVEN OTHER enz’s to catalyze lysis of modulator • So have groups of enzymes • Not necessarily subunits that interact

  25. Covalent Modification at Reg Enz Funct’l Grps • Could disrupt entire 2o, 3o structure • Could inhibit S approach • Could inhibit S fit • Could modify funct’l grps impt to catalysis

  26. Types of Modification • Phosphorylation Nucleotidation • ADP-ribosylation Methylation/acetylation

  27. Glycogen Phosphorylase Example • Glycolysis reg enz • 2 subunits, each w/ ser (what’s it’s funct’l grp?) • Cleaves glycogen (what’s it made of?) • Releases glu then phosphorylates glu

  28. 2 forms of enz (a = active, b = inactive) • 2 assoc’d enz’s • Phosphorylase kinase cat’s b  a • Active form – phosphorylated • W/ transfer of Pi from ATP • Phosphorylase phosphatase cat’s a  b • W/ hydrolysis Pi

  29. Phosph’n interferes w/ stabilizing ionic interactions • Changes folding (what type of structure (1o, etc.) is most impt to folding?) • New interactions between diff aa’s • Incr’s catalytic activity • a, b forms differ in 2o, 3o, 4o structures also • So some allosteric properties

  30. Another Kinase May Activate Glycogen Phosphorylase • Protein kinase A is modulated also • Works through 2nd messenger system:

  31. Phosphorylation & Second Messenger Systems • Involves both allosteric & cov’ly mod’d enzymes • “First messenger” = non-lipid hormone • Binds cell membr receptor • Book ex: epinephrine

  32.  Conform’l changes in membr-bound proteins • Receptor, G-proteins, adenylyl cyclase • All are allosteric prot’s

More Related