Apple Experiment

1. Apple Experiment Come down and get an apple and a slice of lemon. When you get back to your seat: 1. Take a big bite of your delicious apple. 2. Immediately squeeze lemon juice over the apple flesh that is now exposed from the bite. 3. IMPORTANT! Don’t get lemon juice all over the apple. Make sure that it is ONLY on the area that you just bit! 4. Set the lemon aside and wipe any lemon juice off of your hands with a napkin. 5. Take another bite from the opposite side of your apple. 6. Set your apple aside.

2. What Are Enzymes? • Enzymes are __________(tertiary and quaternary structures). • An enzyme is a _________ in cellular reactions. • A catalyst accelerates a reaction.

3. Enzymes • Are ________ for what they will catalyze • Are _________ • Usually end in -_____ -Sucrase -Lactase -Maltase

4. How do enzymes work? • Each enzyme has a unique 3-D shape, including a surface groove called an ______ _____. • The enzyme works by binding a specific chemical reactant (_________) to its active site, causing the substrate to become unstable and react. • The resulting __________ is then released from the active site.

5. How do enzymes work? Enzymes work by _______ ______ which ________activation energy.

6. Enzyme Enzyme-Substrate Complex The reactant an enzyme acts on is specifically called the _________. Joins Substrate

7. Enzyme • A specific regionof an enzyme molecule which binds to the substrate. Active Site Substrate ______ _____

8. An enzyme fits with its substrate like a ____ and ____. Shape of a Protein

9. EnzymeAnimation : Gary E. Kaiser http://student.ccbcmd.edu/biotutorials/proteins/enzsub.htmlView : http://highered.mcgraw-hill.com/sites/0072495855/student_view0/chapter2/animation__how_enzymes_work.html

10. Enzymes are ____ ________ in the reactions they catalyze.Think of them as tiny machines in manufacturing. The more machines, the faster the accumulation of _________. Image: Wine Bottling : www.morrison-chs.com/timingscrews/index.html Wine Vats: www.lymebaywinery.co.uk/pages/about_us.htm

11. ( ________ ) _______ + ________ -----------> _________ ( ________ ) __________ -----------> ________ ________ Formats for writing a chemical reaction.

12. 4 levels of Protein Structure

13. How Do You Stop an Enzyme? _______________ • Alteration of a protein shape through some form of external stress • Denatured protein can’t carry out its cellular function . Irreversible egg protein denaturation caused by high temperature (while cooking it).

14. Temperature • pH • Cofactors & Coenzymes • Inhibitors Factors Affecting Enzyme Activity

15. Temperature & pH • Think about what kind of cell or organism an enzyme may work in… • Temperatures far above the normal range _________ enzymes • (This is why very high fevers are so dangerous. They can cook the body’s proteins) • Most enzymes work best near __________ pH (6 to 8).

16. Coenzymes & Cofactors • Non-protein substances (zinc, iron, copper, vitamins) are sometimes need for proper enzymatic activity. • Coenzyme versus Cofactor: What’s the dif? _________ more general term. Includes inorganic and organic molecules. _________ type of Cofactor, But specifically organic molecules. Image: EnzymeCofactor : Public domain Wiki, Ribbon-diagram showing carbonic anhydrase II. The grey sphere is the zinc cofactor in the active site.

17. Coenzyme : Vitamin B12 • Example: Most _________ are coenzymes essential in helping move atoms between molecules in the formation of carbohydrates, fats, and proteins. • Exclusively synthesized by ___________ (found primarily in meat, eggs and dairy products). Image: VitaminB12 : NIH, Public Domain www.nlm.nih.gov/.../ency/imagepages/19516.htm

18. Enzyme Competitive inhibitor • ___________ ____________: Chemicals that resemble an enzyme’s normal substrate and compete with it for the active site. Substrate Two Types of Enzyme Inhibitors

19. 1. Competitive inhibitors: Resemble an enzyme’s normal substrate and compete with it for the active site. Image: Competitive Inhibition : www-biol.paisley.ac.uk/.../chapter3_2.html

20. _______________ ______________: Do not enter the active site, but bind to another part of the enzyme causing the enzyme to change its shape, altering the activesite. Noncompetitive Inhibitor Enzyme active site altered Substrate Two Types of Enzyme Inhibitors

21. Enzyme Inhibitors Blocking an enzyme's activity can kill a pathogen or correct a metabolic imbalance. Many _____ are enzyme inhibitors. Enzyme inhibitors are also used as _________ and __________. Images Dead Bug : www.kansas.gov/help_center/user_testing.html Prescription Drugs : www.patentdocs.us/.../08/by-kevin-e-noon.html

22. Regulation of Enzyme Activity

23. (1) Allosteric control • Covalent modification *changes in enzyme levels due to regulation of protein synthesis or degradation are additional, long-term ways to regulate enzyme activity Enzyme activity is regulated by four different mechanisms*

24. (1) Allosteric regulation = the activation or inhibition of an enzyme’s activity due to binding of an effector molecule at a regulatory site that is distinct from the active site of the enzyme (2) Allosteric regulators generally act by increasing or decreasing the enzyme’s affinity for the substrate Allosteric regulation of enzyme activity

25. (1) Covalent modification allows an enzyme to be rapidly activated or inactivated (2) With covalent modification, regulation of a enzyme activity is achieved at low energy costs to the cell (i.e. regulation does not require synthesis of a new enzyme or inhibitory protein). (3) Phosphorylation is a good example of how enzymes are activated and inactivated by covalent post-translational modifications Covalent modification

26. Can either activate it or inhibit it by altering the conformation of the enzyme or by serving as a functional group in the active site. Covalent modification regulates the catalytic activity of some enzymes

27. (1) Allosteric regulation ATP activation/CTP inhibition of ATCase sigmoidal kinetics cAMP activation of cAMP-dependent protein kinase (2) Reversible covalent modification Phosphorylation Ser/Thr protein kinases, Tyr kinases, kinase cascades Summary of regulatory mechanisms

28. Allosteric Enzyme ATCase O H2N-C-O-PO32- = COO- CH2 HN-C-COO- H H COO- CH2 N-C-COO- H H - - O H2N-C- - = - - - - - ATP Feedback inhibition CTP CTP CTP CTP CTP CTP CTP Nucleic acid metabolism Active relaxed form Carbamoyl phosphate Aspartate Carbamoyl aspartate + ATCase Quaternary structure CCC Catalytic subunits R R R R R R Regulatory subunits CCC Catalytic subunits Inactive tense form Juang RH (2004) BCbasics

29. Sigmoidal Curve Effect vo ATP CTP vo Noncooperative (Hyperbolic) Positive effector (ATP) brings sigmoidal curve back to hyperbolic Negative effector (CTP) keeps Sigmoidal curve Cooperative (Sigmoidal) Exaggeration of sigmoidal curve yields a drastic zigzag line that shows the On/Off point clearly Consequently, Allosteric enzyme can sense the concentration of the environment and adjust its activity Off On [Substrate] Juang RH (2004) BCbasics

30. Mechanism and Example of Allosteric Effect Kinetics Models Cooperation vo (+) [S] (+) vo (+) [S] vo (-) (-) [S] Allosteric site R = Relax (active) Homotropic (+) Concerted Allosteric site A Heterotropic (+) Sequential X T = Tense (inactive) I Heterotropic (-) Concerted X X Juang RH (2004) BCbasics

31. Activity Regulation of Glycogen Phosphorylase A A P P P P A A A A P P P P P P A A Covalent modification GP kinase T P T P GP phosphatase 1 Non-covalent ATP Glc-6-P Glucose Caffeine spontaneously A Glucose Caffeine AMP Garrett & Grisham (1999) Biochemistry (2e) p.679 A R R A