Introduction

1. Introduction Biochem I Biol 3252 / Chem 3251 2008-09-04

2. Course details Welcome! • See course webpage on WebCT for outline, manual, other documents • Full lecture schedule in outline, will try to keep on track • Office CB4018, Office hour Tuesday 9:30-10:30AM, or email me to make an appointment (dlaw@lakeheadu.ca)

3. Lecture topic summary • More detailed list in outline • Sept. 4-9 General intro and intro to metabolism (2 lectures) • Sept. 11-25 Proteins (5 lectures) • Fri. Sept. 26 Biotechnology seminar: bonus Qs on test 1 • Tues. Sept. 30 Test 1: Metabolism and proteins • Oct. 2-14 Lipids (4 lectures) • Thurs. Oct. 16 Test 2: Lipids • Oct. 23-Nov. 6 Carbohydrates (5 lectures) • Thurs. Nov. 6 Test 3: Carbohydrates • Nov. 11-25 Nucleic acids (5 lectures) • Late Nov./ Review for exam • Early Dec.

4. Course details • Lectures: • Tues and Thurs • Sept. 4 to Nov. 25, 2008, 5:30 – 7:00 PMAT2001 • Labs: • CB2050 and 2051 • Sept 10 to Nov 26 • Section F1: Wed 8:30A – 11:30A • Section F2: Wed 11:30A – 2:30P • Read your lakeheadu.ca email regularly to receive class updates

5. Marking scheme • Three term tests worth 15% each (1 h each, written in class) • Tues. Sept. 30 • Thurs. Oct. 16 • Thurs. Nov. 6 • Final in Dec. worth 30% • Lab worth 20% (4 labs @ 5% each) • Lab coordinator: Jarrett Sylvestre (jarrett_sylvestre@yahoo.ca) • TAs: Caroline Cheng (cscheng@lakeheadu.ca) and Chris Edmunds (caedmund@lakeheadu.ca) • Lab check-in next week (Wed. Sept. 10 and Fri. Sept. 12) • You need a lab coat,safety glasses and a lab manual to participate in the labs • safety glasses and lab coat available in bookstore • lab manual is a PDF on the course WebCT site

6. Participation • Worth 5% of final mark • Using the i>clicker electronic voting system—available in the bookstore either alone or bundled with text • Each class will have at least one question related to the material that has a multiple choice answer • Half marks for trying, half marks for getting them right • Throw out marks for lowest 4 days for each student (= 4 absences excused without penalty over the course)

7. i>clicker • Please register your remote prior to the next class (Tuesday) online (www.iclicker.com/registration) • “Student ID” is your email userid (e.g., “dlaw”) • “Clicker ID” found on the back of your remote (e.g.,00A0A00A)

8. Textbook • (also required for Bchem II) • “Biochemistry”, 6th ed., Berg et al. (2006). • Available in bookstore • The textbook has a multimedia site, where you have access to • interactive exercises • animated 3D tutorials • learning tools • See course website for URL With i>clicker: ~$160 in bookstore Note that the 5th edn text was used until 2005 and is still acceptable

9. Statement on academic dishonesty • Makes up part of the Code of Student Behaviour and Disciplinary Procedures • You should be familiar with its content; URL is in the outline • For this course, cheating may occur by: • Conferring with notes or another person in a test or exam; • Handing in a lab report that is plagiarized • Electronically voting on an absent student’s behalf • All of these will result in a mark of zero for that work • Cheating on an exam or repeated cheating will result in a course mark of zero and possible expulsion from the University • If you cheat • You are cheating yourself! • The instructors will catch you

10. Course overview • In this course, we will talk about the four major types of macromolecules and their subunits, synthesis and degradation, and functions • These macromolecules occur in particular places in the cell and fulfill specific functions • Bchm II next term will integrate these concepts into a holistic discussion of some aspects of their metabolism (enzymes, signal transduction, disease)

11. What is biochemistry? Life seems to be a paradox! “Living things are composed of lifeless molecules”— Albert Lehninger, 1982. • Biochemistry allows us to use chemical and physical rules governing individual molecules to predict behaviour of living organisms • Living matter uniquely: • is highly organized(macroscopically, microscopically) • is made of components with distinct functions (from limbs to fatty acids) • can extract, transform, use energy from environment (fight entropy!) • can self-replicate

12. What separates non-living and living? • 18th century (Enlightenment): vitalism, a/k/a “the philosophick mercury” sought by alchemists universe-review.ca • Now seek testable, repeatable experiments to explain natural phenomena (scientific method) • Chemical molecules inside and outside living organisms act according to physical laws • As part of metabolism, biomoleculesalso act as part of the “molecular logic of the living state”

13. Core biochemical concepts • Biochemical unity • The importance of water for life: interaction with other molecules • Acids and bases: how molecules interact chemically • Buffers and pH: controlling the environment in the cell (homeostasis) • The interdependence of biochemical pathways

14. Core concept 1: Biochemical unity http://www.bio.davidson.edu/courses/Molbio/MolStudents/spring2000/deeb/macdnasis.html • Biomolecules: so different, so identical • 107 species x 50,000 proteins per species = ~1011 proteins! All are different but • Many are similar (homologous) in sequence and function between species (e.g., enzymes such as pyruvate kinase) • They are all made up of the same building blocks: 20 amino acids • From immense simplicity comes immense diversity! Multiple sequence alignment of pyruvate kinase proteins from different kingdoms

15. Biochemical unity suggests a common ancestor • Extends to other macromolecules as well (DNA, RNA, lipids, CHO) and their monomers: identical or extremely similar between species • Has phylogenetic implications: what constitutes a species? • “Species” traditionally defined in Linnean systematics as similar looking (also, can interbreed, etc.) • Though macromolecules of inheritance and function are homologous between species, between individuals within a species (and even within families) they are very close to identical All these organisms have DNA, RNA, proteins constructed the same way! Alberts et al., Molecular biology of the cell, 4th edn.

16. Similarity extends to the protein level as well • Look at the shape of the same regulatory protein in species from 3 kingdoms/domains

17. Common ancestry reflected in macromolecule sequences Domain (or Kingdom) • Evident when constructing phylogenetic trees (a/k/a c_____) relating macromolecule (e.g., DNA) sequence to species origins • Usually for one highly conserved gene (e.g., 18S ribosomal RNA gene) Genus now close Related genuses/ families/ phyla Time Fig. 1.3 MYA distant Common ancestor • Biochemical unity is reduced the older the split between species but core similarities remain predominant • Tree of life is being reformatted based on biochemical information!

18. Life = using energy for useful work • Harness energy from metabolism to do useful work (Gibbs free energy, ΔGo’) • Common energy currency in all living organisms Useful energy Dissipated energy • Living organisms not in equilibrium with environment: fight entropy, “selfishly” store energy in anabolism by making useful molecules Fig. 1.3, Lehninger, Principles of biochemistry (1982). • Energy is conserved & stored through the formation of chemical bonds: the chemistry of molecules important to life • Let’s walk through a brief review of chemical bonds of importance in biochemistry

19. Chemical bond types Uncredited structural formulas are from text, 5th ed. Covalent • Formed by sharing outer valence shell electrons to fill shell • One shared e- pair = 1 (single) bond • Two shared pairs = double bond, etc. • We can talk about bond energy • C-C is 346 kJ/mol These can form in biological molecules such as purine bases • Strong bonds result if e- can form resonance structures, e.g., benzene (C6H6; C=C is 602 kJ/mol); carbonyl (C=O ~800 kJ/mol) p. 7 of text • Chemical reactions break and form covalent bonds e.g., nucleophilic attack forms new covalent bond between molecules

20. Noncovalent bonds • depend on dipole-dipole interactions • recall that a dipole is an object whose centers of positive and negative c______ do not coincide • this occurs because many molecules possess electron-rich and –poor regions • these regions can interact and result in noncovalent bonding between molecules • e.g., HCl: electrically neutral but possesses a dipole H – Cl • e.g., H2O

21. Noncovalent chemical bond types Electrostatic • Dependent on electrical charges between noncovalently bound atoms • According to Coulomb’s Law: E = k q1 q2 / r2 • a/k/a ionic bonds: a chemical bond in which one atom loses an electron to form a positive ion and the other atom gains an electron to form a negative ion • Example: NaCl composed of two ions: Na+ (e- donor) and Cl- (e- acceptor) Closer = higher energy p. 7 of text Forms tightly packed lattice http://cwx.prenhall.com/bookbind/pubbooks/hillchem3/medialib/media_portfolio/text_images/CH02/FG02_09.JPG

22. Electrostatic interactions also guide the shape of DNA • PO42- (a/k/a Pi) groups in DNA • Form unfavorable electrostatic interactions over distances • Oppose dsDNA formation • BUT: water has a high d_____ constant (or relative static permittivity), meaning that it is very p______ • Also, in vivo, there are also many mono- and divalent cations present (e.g., ____?) • These help neutralize the negative charge of Pi groups and thus permit their presence in DNA

23. Noncovalent chemical bond types Hydrogen • Relatively weak (4-12 kJ/mol) but are crucial for three-dimensional structure of biomolecules (NAs, protein) • Of primary importance in biochemistry: water can H-bond with itself or can easily interact with other molecules: “universal solvent” Hydrogen donor Hydrogen acceptor Water swaps its H-bonds with those between other molecules Example: H-bonds between complementary bases in DNA http://www.biosci.ohiou.edu/introbioslab/Bios170/170_8/at.html

24. Noncovalent chemical bond types Attractive from a distance… …but repellent close up Van der Waals • A force acting between nonbonded atoms or molecules • Relatively weak, even smaller than hydrogen bonds (2-4 kJ/mol) • Based on the fact that charge distribution around atoms is dynamic (e- cloud constantly in motion) • Differences in one atom’s charge distribution complementarily perturb neighboring distributions • These asymmetries attract • Example: attraction between phenyl groups in neighboring phenylalanine residues in a protein • Play major role in protein folding Fig 1.10

25. Van der Waals contacts also mediate the shape of DNA • Another example of biologically important noncovalent bonds promoting formation of complex biomolecules • Bases within the dsDNA interact at the optimum distance to attract each other • Temporary dipoles form that can attract atoms transiently

26. Things to remember about noncovalent bonds • More than one type of noncovalent interaction can occur at once • Though individual bond strengths are small, summed over entire biomolecules they are chemically (and biologically!) significant DNA protein http://www.nature.com/emboj/journal/v19/n4/full/7592187a.html

27. All types of noncovalent bonds regulate how biomacromolecules interact • Weak, noncovalent interactions seem less important than covalent bonds but are crucial biochemically • Examples: • Substrate-enzyme • Hormone-receptor • Protein-protein Substrate attracted to Enzyme via one or combination of noncovalent interactions Image: http://www.chemistry.wustl.edu/~edudev/LabTutorials/Carboxypeptidase/carboxypeptidase.html

28. Core concept 2: Water is essential for life • In biochemistry, noncovalent interactions rely on the properties of water • Polar: asymmetrically distributed charge: bent molecule! • H-bonds hold water together cohesively • Universal solvent: disrupts and weakens electrostatic interactions, forms solvent shells around ions, dissolves essential polar molecules so that they can move/diffuse inside cell • Presents a problem for crucial interactions between polar molecules • Hydrophobic microenvironments present in the cell • So many molecules to dissolve, so little water: water is limiting inside the cell

29. Core concept 3: acids and bases • Water dissolves compounds • These are often charged in solution • pH-dependent • Depends on shape & atoms in molecule: many are non-planar • Can become • protonated = basic (e.g., ammonia: NH3) • deprotonated = acidic (e.g., acetate: CH3COOH) • Thus an acid is a proton donor, and a base is a proton acceptor HX ↔ H+ + X- Note that acids and bases are not charged! conjugate base acid conjugate acid base Image: http://www.windows.ucar.edu/tour/link=/physical_science/chemistry/ammonia.html

30. Core concept 3: acids and bases • Water itself dissociates into ions: • We can quantify how readily this dissociation happens by calculating a dissociation constant (Keq): • Water is mostly nonionized: Keq at 25°C is 10-14 • We can also calculate Keq for other species too, like acids • Recall that pH is a measure of the H+ concentration of a solution: • We can similarly define pK for an acid: Conc products = Large (>>1): mostly products Small (1<Keq<0): mostly substrates Conc reactants , so

31. Core concept 3: acids and bases • pH and pK are related (Henderson-Hasselblach equation): • If an acid HA is half dissociated to H+ and A-, then [A-] = [HA] • Thus, pK is the pH at which half of the acid is dissociated • This is important when considering the buffering capacity of these molecules in solution, which leads us to… can be rearranged to give Then = 1, log = 0, and pH = pK.

32. Core concept 4: buffers and cellular pH • Acid-base conjugate pairs (e.g., acetic acid and acetate) in solution resist changes in the solution’s pH: they are buffers • Consider adding base (OH-) to acetic acid (HA): • Remember titration curves from analytical chem? • Titrate OH- into a solution of acetic acid to force its dissociation: Add more OH-, pH does not change as radically Minimum change in centre of curve where [A-] = [HA] This is the pK of acetic acid! 2 SIGMOIDAL RESPONSE! Initially, small changes in [OH-] produce large changes in pH 3 3 1 2 1 Stryer, “Biochemistry”, 3rd edn. (1988)

33. Core concept 4: buffers and pH What does this chemistry have to do with biochemistry? • Weak acids are effective at buffering their environment at pH values near their pK • pK values indicate at what pH a functional group is ionized (titrated) a proton from a molecule • It can take a small amount of OH- (at low pH): e.g., -COOH  -COO- (pH 2.3) • Or a high amount of OH- (at high pH): e.g., -NH3+ -NH2(pH 9.6) Stryer, “Biochemistry”, 3rd edn. (1988) Increasing pH Note: There are many carboxyl and amine groups in biomacromolecules!

34. Core concept 4: buffers and pH • Protein amino acid sidechains have different R (functional) groups, each with their own pK • Proteins are more or less ionizable depending on • pH of environment • Types of R groups they contain • This affects • how they interact with other proteins • activity • solubility • Natural (in vivo) buffers include the bicarbonate ion • Cells must control their interior pH (of their intracellular space, or c________) • Proteins, enzymes, have optimal pH for their activity • The core concepts we covered are crucial for understanding how molecules are made and interact (e.g., for the rest of this course!)

35. Core concept 5: the interdependence of biochemical pathways And finally! • Though the course will present metabolism of the four macromolecule types as largely distinct, all metabolic pathways are linked • Share products • Macromolecules combine to change their function, e.g., • Proteins decorated with lipids, carbohydrates • Complexes of nucleic acids and proteins • This will be addressed in greater detail when we discuss the integration of metabolism Fig 15.2