Advanced Biochemistry BISC 6310 Fall 2011

1. Advanced Biochemistry BISC 6310 Fall 2011

2. Course topics:Textbook: Biochemistry, 5th ed. (LubertStryer) • To be tought by Dr. Tarek M. Zaida… • Basic concepts of metabolism (ch. 8 & 14) • Glycolysis and gluconeogenesis (ch. 16) • The citric acid cycle (ch.17) • Oxidative phosphorylation (ch. 18) • Glycogen metabolism (ch. 21) • Fatty acids metabolism (ch. 22)

3. To be tought by Dr. KamalAlkahlout… 7. Protein turnover and amino acids metabolism (ch. 23) 8. Synthesizing the molecules of life: 9. Biosynthesis of Amino acids (Ch.24) 10. Nucleotide biosynthesis (ch.25) 11. Biosynthesis of membrane lipids & steroids (ch. 26) 12. The Integration of metabolism (ch. 30) 13. Signal Transduction (ch.15) 14. The Calvin cycle (ch.20) 15. Biochemical Techniques

4. Activities and exams • Activities: • At the end of the term every student will present an oral presentation of their choosing, covering one of the medical-related disorders connected to any of the course chapters • Note: You have the right to choose a topic, but you are obligated to maintain your focus on the biochemical background of the topic. • You have to let us know which topic you have chosen within 2 weeks from now, so that we may grant you the permission to proceed with your research.

5. The evaluation of the presentation will be based on: • Performance (oral) • Content (oral & written) • Time frame (limited to 10 min.) • Note: Be ready to give me up your presentation electronically or via email at : tzaida@iugaza.edu or tarekzaida@yahoo.com • The time of the presentation should not exceed 10 minutes. • Exams: • Two exams will be held in the middle and at end of the semester. • The material of the final exam will include up to 20% of the chapters I will cover, the other 80% of the final exam will be covered by the chapters tought by Dr. Kamal.

6. You will be given 2 Quizzes., the 1st before midterm & the 2nd before final exam. • Making-ups are not allowed neither for presentations nor for quizzes nor for midterm ……………(STRICTLY ENFORCED) • Class attendance is strongly recommended, But once you decide not to come to class, its your responsibility to keep yourself posted and updated with what has been done/ said during class. • I will always welcome your suggestions as it is the best way to improve the outcome of this class(you can talk to me in person or if you feel more comfortable drop me an email) • Office hours: TBD

7. Grading:

8. METABOLISMBASIC CONCEPTS & DESIGN The quantitative study of cellular energy transductions and the chemical reactions underlying these transductions.

9. Enzymes • Enzymesare the most sophisticated biological catalysts known. • catalysts alter the rates of chemical reactions but are neither formed nor consumed during the reactions they catalyze. • Most enzymes are proteins. Some nucleic acids exhibit enzymatic activities (e.g., rRNA).

10. Enzyme Characteristics:Rate Enhancement • Enzymes significantly enhance the rates of reactions:

11. Enzyme Characteristics:Turnover Numbers • The “turnover number” is used to rate the effeciency of an enzyme. • It tells how many molecules of reactant a molecule of enzyme can convert to product(s) per second. • How fast can an enzyme produce products?

12. Enzyme Characteristics:Specificity • Enzymes can be very specific. • For example, proteolytic enzymes help hydrolyze peptide bonds in proteins. • Trypsin is rather specific • Thrombin is very specific The specificity of an enzyme is due to the precise interaction of substrate with the enzyme. This is a result of the intricate three-dimensional structure of the enzyme protein.

13. Enzyme Characteristics:Regulation • Enzymes can enhance the rates of reactions by many orders of magnitude. • A rate enhancement of 1017 means that what would occur in 1 second with an enzyme’s help, would otherwise require 31,710,000,000 years to take place. • Thus, regulation of enzymatic activity is in a sense, regulation of metabolism, or any other cellular process

14. Enzyme Cofactors • Many enzymes use same cofactor • Cofactors are split into two groups: • Metals • Coenzymes (small organic molecules) • Most vitamins are coenzymes. • When tightly bound to enzyme, cofactor =prosthetic group “Apoenzyme” + cofactor = “Holoenzyme”

15. Enzymes – Gibbs Free Energy • Gibb’s “Free Energy,” ΔG, determines the spontaneity of a reaction: • ΔG must be negative for a reaction to occur spontaneously • A system is at equilibrium and no net change can occur if ΔG is zero • A reaction will not occur spontaneously if ΔG is positive; to proceed, it must receive an input of free energy from another source.

16. G = Hsystem - TSsystem • DG is the Gibbs free energychange, used to describe energetics of biochemical reactions • DH is the change in enthalpy,heat content of a system. • DS is the change in entropy, a measure of the level of randomness or disorder in a system. • The total entropy of a system and its surroundings always increases for a spontaneous process) {2nd Law} • Entropy will increase only if: G = Hsystem - TSsystem < 0 • The free-energy change must be negative for a reaction to be spontaneous

17. G of a reaction depends only on free-energy of products minus free-energy of reactants. • G of a reaction is independent of path (or molecular mechanism) of the transformation • G provides no information about the rate of a reaction • For the reaction: A + B  C + D • To determine G, must consider nature of both reactants and products as well as their concentrations

18. Free energy & equilibrium constant (K) • At equilibrium, G = 0. • 0 = Go’ + RTln([C][D]/[A][B]) • K’eq =[C][D]/[A][B] • Go’ = - RTlnK’eq • An enzyme cannot alter the equilibrium of a chemical reaction. • This means, an enzyme accelerates the forward and reverse reactions by precisely the same factor.

22. Enzymes decrease activation energy • A chemical reaction goes through a transition state with a higher G than either S of P • Enzymes facilitate the formation of the transition state by decreasing G‡

23. Enzyme-Substrate Complex • For enzymes to function, they must come in contact with the substrate. • While in contact, they are referred to as the “enzyme-substrate complex.” • The combination of substrate and enzyme creates a new reaction pathway, with a lowered transition-state energy • More molecules have the required energy to reach the transition state • Catalytic power of enzymes is derived from the formation of the transition states in enzyme-substrate (ES) complexes • The essence of catalysis is specific binding of the transition state

24. Enzyme – Active Site • Enzymes are often quite large compared to their substrates. The relatively small region where the substrate binds and catalysis takes place is called the “active site.” (e.g., human carbonic anhydrase:)

25. Enzyme – Active Site • The active site is the region that binds the substrates (& cofactors if any) • It contains the residues that directly participate in the making & breaking of bonds (these residues are called catalytic groups) • The interaction of the enzyme and substrate at the active site promotes the formation of the transition state • The active site is the region that most directly lowers G‡ of the reaction - resulting in rate enhancement of the reaction

26. Enzymes differ widely in, structure, specificity, & mode of catalysis, yet, active site have common features: The active site is a 3-D cleft formed by groups that come from different parts of the amino acid sequence Water is usually excluded unless it is a reactant. Substrates bind to enzymes by multiple weak attractions (electrostatic interactions, hydrogen bonds, hydrophobic interactions, etc. The specificity of binding depends on the precisely defined arrangement of atoms at the active site

27. Enzyme Classification • Enzymes are classified and named according to the types of reactions they catalyze: • Proteolyticenzymes [such astrypsin] lyse protein peptide bonds. • “ATPase” breaks down ATP • “ATP synthetase” synthesizes ATP • “Lactate dehydrogenase” oxidizes lactate, removing two hydrogen atoms. • Such a wide variety of names can be confusing. A better method was needed.

28. Enzyme Classification • The “Enzyme Commission” invented a systematic numbering system for enzymes based upon these categories, with extensions for various subgroups. e.g., nucleoside monophosphate kinase (transfers phosphates) • EC 2.7.4.4. 2 = Transferase, 7 = phosphate transferred, 4=transferred to another phosphate, 4 = acceptor

29. LIVING ORGANISMS NEED ENERGY FOR: • Performing mechanical work • Active transport and maintaining homeostasis • Synthesis of macromolecuels and biochemicals.

30. METABOLIC PATHWAYS • Catabolic pathways • Anabolic pathways • Amphibolic pathways that can be both anabolic and catabolic depending on the energy status in the cell.

31. The useful forms of energy produced in catabolism are employed in anabolism

32. A METABOLIC PATHWAY MUST SATISFY MINIMALLY TWO CRITERIA • The reactions of the pathway must be specific: • This criterion is accomplished by enzymes • The entire set of reactions must be thermodynamically favored. • A reaction can occur spontaneously only if DG, the change in free energy, is negative.

33. AN IMPORTANT THERMODYNAMIC FACT • The overall free-energy change for a chemically coupled series of reactions is equal to the sum of the free energy changes of the individual steps.

34. The two reactions are catalyzed by the same enzyme

40. ATP IS THE ENERGY CURRENCY IN BIOLOGICAL SYSTEMS The active form of ATP is usually attached to Mg2+ or Mn2+

41. ATP IS THE ENERGY CURRENCY IN BIOLOGICAL SYSTEMS • ATP is an energy-rich molecule because its triphosphate unit contains two phosphoanhydride bonds. • A large amount of free energy is liberated when ATP is hydrolyzed to ADP and (Pi) or to AMP and PPi.

42. ATP IS THE ENERGY CURRENCY IN BIOLOGICAL SYSTEMS • The precise DG°’ for these reactions depends on: • The ionic strength of the medium • The concentrations of Mg2+ and other metal ions. • Under typical cellular concentrations, the actual DG for these hydrolyses is approximately -12 kcal mol-1 (-50 kJ/mol). • ATP hydrolysis can be coupled to promote unfavorable reactions.

43. ATP Hydrolysis Drives Metabolism by Shifting the Equilibrium of Coupled Reactions

44. A can be converted into B if the reaction is coupled to ATP hydrolysis

45. ATP is an energy coupling agent: The hydrolysis of an ATP molecule in a coupled reaction changes the equilibrium by a factor of 108 (1.15 X10-3 compared to 1.34X105)

46. A and B in the preceding example can be any two different chemical species • A and B may represent activated and unactivated conformations of a protein • phosphorylation with ATP may be a means of conversion into an activated conformation. • Such a conformation can store free energy, which can then be used to drive a thermodynamically unfavorable reaction. (muscle contraction). • A and B may refer to the concentrations of an ion or molecule on the outside and inside of a cell, as in the active transport of a nutrient.

47. The Structural Basis Of The High Phosphoryl Transfer Potential Of ATP • Resonance stabilization: • ADP and, particularly, Pi, have greater resonance stabilization than does ATP. • Electrostatic repulsion: • At pH 7, the triphosphate unit of ATP carries about four negative charges in close proximity. • The repulsion between them is reduced when ATP is hydrolyzed. • Stabilization due to hydration: • Water can bind more effectively to ADP and Pi than it can to the phosphoanhydride part of ATP, stabilizing the ADP and Pi by hydration.

48. Phosphoryl Transfer Potential

49. Phosphoryl Transfer Potential • It is significant that ATP has an intermediate phosphoryl transfer potential among the biologically important phosphorylated molecules. • This intermediate position enables ATP to function efficiently as a carrier of phosphoryl groups. • If ATP had the highest potential then it wouldn’t be formed.