BCH 201 General Biochemistry I Lecture One PDF

BCH 201: General Biochemistry I Dr. Adedeji Nelson Ademakinwa (Deanry, Room 46) |Floor One) Venue: PFA 9-11|Tuesday.  Course Outline WEEK 1 Introduction Dr. Ademakinwa A.N. 2 Proteins and the...

BCH 201: General Biochemistry I Dr. Adedeji Nelson Ademakinwa (Deanry, Room 46) |Floor One) Venue: PFA 9-11|Tuesday.  Course Outline WEEK 1 Introduction Dr. Ademakinwa A.N. 2 Proteins and their Functions Dr. Ademakinwa A.N. 3 Chemistry of Amino Acids Dr. Ademakinwa A.N. 4 Protein Structure Dr. Ademakinwa A.N. 5 Enzymes and co-enzymes Dr. Ademakinwa A.N. 6 Structures of carbohydrates Dr. Ademakinwa A.N. 7 Structure of nucleic acids Dr. Ademakinwa A.N. 8 Effects of alkali and acids on nucleic acids Dr. Famurewa A.J. 9 Buffers: Chemistry (acidity & alkanility) Dr. Famurewa A.J. 10 Buffers: pH and pKa values (Calculations) Dr. Famurewa A.J. 11 Methods of studying metabolism Dr. Famurewa A.J.  Proteins Proteins are naturally-occurring biopolymers comprised of amino acids. The biological function of proteins is inherent in their three dimensional structure. All the information required for correct folding of the protein into its functional native structure is contained in the primary sequence of amino acids. The physical chemical properties of the amino acids contain the biological information required for folding and function.  Protein - Daily Requirements Average adult contains ~10kg of protein;~300g is replaced daily by recycling and intake. We need to take in ~70g of high quality protein or ~80g of lower quality this varies with age, size and energy demand,eg. infants: 1.8g/kg/day children: 1.0g/kg/day adults: 0.8g/kg/day Recommended: ~15% of daily Caloric intake Normally the body does not store proteins. Since they are the major source of nitrogen they are constantly being broken down and reconstructed. Protein is lost in urine, fecal material, sweat, hair/nail cuttings and sloughed skin.  Proteins: Biological Function Catalysis – enolase (in the glycolytic pathway) – DNA polymerase (in DNA replication) Transport – hemoglobin (transports O2 in the blood) – lactose permease (transports lactose across the cell membrane) Structure – collagen (connective tissue) – keratin (hair, nails, feathers, horns) Motion – myosin (muscle tissue) – actin (muscle tissue, cell motility)  Proteins by Structure Proteins Simple Conjugated Fibrous Globular Lipo- Glyco- Hemo- insoluble soluble ‘structural’ ‘reactive’ hair, horn enzymes HDL, interferon hemo- LDL globin  Proteins by Structure Fibrous Collagens Elastins Keratins Myosins bones lungs hair/feathers muscles tendons ligaments horn/nails cartilage  Proteins by Structure Globular Albumins Globulins egg whites antibodies(-globulin) enzymes  Amino Acids: Building Blocks of Protein Proteins are linear heteropolymers of -amino acids. Amino acids have properties that are well suited to carry out a variety of biological functions: capacity to polymerize useful acid-base properties varied physical properties varied chemical functionality  (Non)Essential Amino Acids The essential amino acids (10) are those that our bodies cannot synthesize. We must obtain them from our dietary intake. They are: histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, valine (and arginine in infants). The non-essential a.a.(10) can be synthesized in our bodies from breakdown products of metabolism. Amino Acids 1. Alanine 1. Isoleucine 2. Asparagine 3. Aspartic acid 2. Leucine 4. Cysteine 3. Lysine 5. Glutamic acid 4. Methionine 6. Glutamine 5. Phenylalanine 7. Glycine 8. Proline 6. Threonine 9. Selenocysteine 7. Tryptophan 10. Serine 8. Valine 11. Tyrosine 12. Arginine 9. Histidine 13. Ornithine  Amino Acids - Protein building blocks An amino acid is a compound having both a carboxyl group(-COOH) and an amino group(-NH2). H All amino acids from protein have the -NH2 attached at H 2N C COOH the C  to the –COOH R (as well as the H- & R-). All naturally occurring -amino acids, except glycine (R=H), are chiral and the ‘L’ stereoisomer.  H H 2N C COOH There are 20 -amino acids R in naturally occurring protein. By convention the -NH2 is placed ‘to the left’. Each aa has a ‘common’ name often ending in ‘-ine’. There are ~150 other physiologically important amino acids, GABA (a neurotransmitter). Amino Acids You must know: Their names Their structure Their three letter code Their one letter code O H 2N CH C OH CH2 Tyrosine, Tyr, Y, aromatic, hydroxyl OH Amino Acids Have Three Common Functional Groups Attached to the α Carbon The α carbon always has four substituents and is tetrahedral. All (except proline) have: an acidic carboxyl group connected to the α carbon a basic amino group connected to the α carbon an α hydrogen connected to the α carbon The fourth substituent (R) is unique in glycine, the simplest amino acid. The fourth substituent is also hydrogen.  20 Amino acids Glycine (G) Alanine (A) Valine (V) Isoleucine (I) Leucine (L) Proline (P) Methionine (M) Phenylalanine (F) Tryptophan (W) Asparagine (N) Glutamine (Q) Serine (S) Threonine (T) Tyrosine (Y) Cysteine (C) Asparatic acid (D) Glutamic acid (E) Lysine (K) Arginine (R) Histidine (H) White: Hydrophobic, Green: Hydrophilic, Red: Acidic, Blue: Basic Amino acids vary in Size Structure Electric charge Solubility in water  Amino Acids: Classification Common amino acids can be placed in five basic groups depending on their R substituents: nonpolar, aliphatic (7) aromatic (3) polar, uncharged (5) positively charged (3) negatively charged (2) Aliphatic Amino Acids Aromatic Amino Acids Polar Amino Acids Acidic Amino Acids Basic Amino Acids Amino acids and absorption  Amino acids are Ampholytes They can act as either an acid or a base They are Zwitterions or molecules that have both a positive and a negative charge Because of their ionic nature they have extremely high melting temperatures  Acid - Base properties of amino acids  [A - ]  pH = pK + log   [HA]  Isoelectric point: the pH where a protein carries no net electrical charge pI = (pK i + pK j ) 1 2 For a mono amino-mono carboxylic residue pKi = pK1 and pKj = pK2 ; for D and E, pKi = pK1 and pKj - pKR ; For R, H and K, pKi = KR and pKj = pK2  Ionization of Amino Acids Amino acids contain at least two ionizable protons, each with its own pKa. The carboxylic acid has an acidic pKa and will be protonated at an acidic (low) pH: −COOH COO− + H+ The amino group has a basic pKa and will be protonated until basic pH (high) is achieved: −NH4+ NH3 + H+  Ionization of Amino Acids At low pH, the amino acid exists in a positively charged form (cation). At high pH, the amino acid exists in a negatively charged form (anion). Between the pKa for each group, the amino acid exists in a zwitterion form, in which a single molecule has both a positive and negative charge.  Acid-Base Behavior of Amino Acids Some amino acids carry a prototropic side chain e.g, aspartic acid have extra carboxyl, histidine has extra imidazole, lysine has an amino and arginine has a guadino group. The pKa values of some amino acids are indicated For amino acids, the various groups have a charge that depends on the pH of the solution The protonated forms of the carboxyl groups and the tyrosine side chain are uncharged while the deprotonated forms are negatively charged or anionic The protonated forms of the amino group, the imidazolium side chain of histidine and the guanidinium group are positively charged while the deprotonated forms are uncharged Charge on Polybasic amino acids and Isolectric point At pH values below the pKa of a group, the solution is more acidic and the protonated form predominates. As the pH is raised above the pKa of a group, that group loses its proton, i.e. the –COOH becomes – COO-, while the positively charged form of the amino group, -NH3+ becomes uncharged, -NH2 As a general rule, the isoelectric point of an amino acid is the average of the pKa values of the protonation transitions on either sides of the isoelectric species; this implies a knowledge of the charge of the various forms Cation → Zwitterion → Anion  Amino Acids Carry a Net Charge of Zero at a Specific pH (the pI) Zwitterions predominate at pH values between the pKa values of the amino and carboxyl groups. For amino acids without ionizable side chains, the Isoelectric Point (equivalence point, pI) is: pK1 + pK 2 pI = 2 At this point, the net charge is zero. – AA is least soluble in water. – AA does not migrate in electric field.  Amino Acids Can Act as Buffers Amino acids with uncharged side chains, such as glycine, have two pKa values: The pKa of the -carboxyl group is 2.34. The pKa of the -amino group is 9.6. As buffers prevent change in pH close to the pKa, glycine can act as a buffer in two pH ranges. Buffer Regions  Ionizable Side Chains Also Have pKa and Act as Buffers Ionizable side chains influence the pI of the amino acid. Ionizable side chains can be also titrated. Titration curves are now more complex, as each pKa has a buffering zone of 2 pH units.  When the Side Chain Is Ionizable Identify species that carries a net zero charge. Identify the pKa value that defines the acid strength of this zwitterion: (pKR). Identify the pKa value that defines the base strength of this zwitterion: (pK2). Take the average of these two pKa values. What is the pI of histidine? Peptide Bonds Link Amino Acids Form when the acid group (COOH) of one amino acid joins with the amine group (NH2) of a second amino acid Formed through condensation Broken through hydrolysis  Amino Acids Polymerize to Form Peptides Peptides are small condensation products of amino acids. They are “small” compared with proteins (Mw < 10 kDa).  Peptide Ends Are Not the Same Numbering (and naming) starts from the amino terminus (N-terminal). AA1 AA2 AA3 AA4 AA5 Condensation and Hydrolytic Reactions Figure 6.3  The tetra peptide Ala-Tyr-Asp-Gly or AYDG Greek lettering used to identify atoms in lysine or glutamate  Amino acids can form peptide bonds Amino acid residue Proteins are molecules that peptide units consist of one or more polypeptide dipeptides chains tripeptides oligopeptides polypeptides Peptides are linear polymers that range from 8 to 4000 amino acid residues There are twenty (20) different naturally occurring amino acids Naming Peptides: Start at the N-terminal Using full amino acid names: serylglycyltyrosylalanylleucine Using the three-letter code abbreviation: Ser-Gly-Tyr-Ala-Leu For longer peptides (like proteins) the one- letter code can be used: SGYAL Assignment A neuropeptide from cockroach as the primary sequence of KLPDPYRREHSWHHTLKVRYYT, illustrate this peptide and determine the net charge at pH 1, 3, 7, 9, 11.  Peptides: A Variety of Functions Hormones and pheromones – insulin (think sugar metabolism) – oxytocin (think childbirth) – sex-peptide (think fruit fly mating) Neuropeptides – substance P (pain mediator) Antibiotics – polymyxin B (for Gram – bacteria) – bacitracin (for Gram + bacteria) Protection, e.g., toxins – amanitin (mushrooms) – conotoxin (cone snails) – chlorotoxin (scorpions) Reaction of amino acids with Ninhydrin  Amino Acid Analysis Acid-hydrolysis of the peptide (6 M HCl, 24 hr) gives a mixture of amino acids. The mixture is separated by ion-exchange chromatography, which depends on the differences in pI among the various amino acids. Amino acids are detected using ninhydrin. Automated method; requires only 10-5 to 10-7 g of peptide.  Question: A peptide was hydrolyzed, and the resulting solution was examined by amino acid analysis, the data Obtained is shown in the table below: Table 1 Solution: Determine the empirical formula Amino acid µmol D 1.21 Amino acid Relative concentration S 0.60 D 2.086 G 1.78 S 1.03 L 0.58 G 3.07 K 0.61 L 1.00 K 1.05 The most likely empirical formula is: 𝐴𝑠𝑝2 , 𝑆𝑒𝑟, 𝐺𝑙𝑦3 , 𝐿𝑒𝑢, 𝐿𝑦𝑠 Assignment: From the empirical formula, illustrate the possible peptide sequence, assuming the first and last Amino acids are Asp and Lys. Determine the net chargeAt pH 1, 3, 5 and 11.