Understanding Protein Structure and Function: Amino Acids, Enzymes, and Protein Folding, Exams of Biochemistry

Download Understanding Protein Structure and Function: Amino Acids, Enzymes, and Protein Folding and more Exams Biochemistry in PDF only on Docsity! Biochemistry 1 Proteins - Polymers of AMINO ACIDS. Residues - another name for AMINO ACIDS Oligopeptide - VERY SMALL chain of amino acids Polypeptide - Longer chain of amino acids Alpha Carbon Stereocenter - All human amino acids are chiral EXCEPT for GLYCINE. They have an R group, a hydrogen, a Carboxylic acid, and an amine. Absolute Configuration - L or D designations are given based on which side of the Fischer Projection an Amine group is. ALL HUMAN AMINO ACIDS ARE L as they are derived from L-Glyceraldehyde All amino acids with the exception of Cysteine (R) and Glycine have an S stereocenter Will a substrate bind in an active site? - Depends on the existence of complementary charges (combining in such a way as to enhance or emphasize the qualities of another-in this case, enhancing the chances of binding and stability) Also depends on the hydrophilicity or basicity How will a protein fold? - Hydrophobic -R groups fold INTO the protein core (hydrophobic environment) Hydrophilic -R groups are more common on the surface of the protein (hydrophilic environment) When you see PROTEIN or ENZYME think: - what amino acids are present and what is the chemistry of their -R groups? pH vs pKa - pKa tells us how acidic (or not) a given hydrogen atom in a molecule is. pH tells us how acidic a solution is. For ex: remember in a titration curve how the pH is the dependent variable (measures the pH of the solution) Pka estimates: alpha -COOH group - pKa ~ 2 Pka estimates: -R Group, ACIDIC - pKA ~ 4 (Asp = 3.7, Glu = 4.5) Pka estimates: -R Group, His - pKa ~ 6 Pka estimates: alpha -NH3+ group - pKa ~ 9 Pka estimates: -R group, BASIC - Peptide Chain Conventions - peptides are WRITTEN, READ, and SYNTHESIZED from N-TERMINUS --> C-TERMINUS Resonance between peptide bonds - resonance between pi electrons of the C=O bond, and the nitrogen pair of the C-N bond, yield 2 resonance structures for any peptide bond. THE ACTUAL STRUCTURE IS A HYBRID of the two. BOTH THE C=O BOND AND THE C-N BOND IN A PEPTIDE BOND HAVE DOUBLE BOND CHARACTER (rigid peptide bond with limited rotation). Trypsin (protein hydrolysis) - cleaves ARGININE and LYSINE on the CARBOXYLIC SIDE think of the one with extra amino groups Chymotrypsin (protein hydrolysis) - cleaves PHENYLALANINE, TRYPTOPHAN, and TYROSINE on the CARBOXYLIC side. think of the R-Groups with aromatic rings It is made of Serine, Histidine, and Aspartate Sulfur Linkage - Gabriel Synthesis - Primary Protein Structure - AA-AA-AA-AA-AA The amino acid sequence Formation of Covalent Peptide Bonds Secondary Protein Structure - Alpha Helices and Beta Pleated Sheets Primarily the result of Hydrogen Bonding Alpha Helices - Hydrogen bonding between CARBONYNL OXYGENS and AMIDE HYDROGENS that are exactly FOUR resides apart. ONLY every FOURTH residue is involved in hydrogen bonding R groups are directed exactly away from the alpha helix cylinder (i.e. perpendicular to a plane tangent to the surface of the alpha helix) Beta Pleated Sheets - Hydrogen bonding between ALL of the CARBONYL OXYGENS in one row and the AMIDE HYDROGENS in the adjacent row. ALL residues are involved in hydrogen bonding R groups are directed perpendicular to the plane of the beta sheet, on both sides. Beta sheets are in a PLEATED conformation. This is necessary for the carbonyl and amide moties to line up properly so that EVERY RESIDUE is participating in TWO HYDROGEN BONDS. Proline - Usually the first residue at the very end of an alpha helix, but rarely found inside the helix because it introduces KINK/TURN. This same KINK/TURN is desirable at the end of beta-sheets because the chain must make a 180 degree turn to align as a neighboring row in a beta sheet. (SEE TOP end of Beta Sheet) Common applications of secondary Structures: Keratin - found in hair and nails = alpha helices Common applications of secondary Structures: Fibroin - molecule that makes up silk = beta sheets Tertiary Protein Structure - Geometric, three-dimensional folding of the alpha helices, beta sheets, and other moieties to form a functional globular or structural protein. 6 Molecular interactions that contribute to a tertiary protein structure - 1) HYDROGEN BONDING between N-H and C=O or side chains (amine, carboxyl, and alcohol groups) 2) DISULFIDE BONDS - covalent bond between 2 cysteine residues 3) HYDROPHILIC/HYDROPHOBIC INTERACTIONS- in soluble proteins, hydrophobic amino acids collapse into the protein core. In membrane proteins, hydrophilic membranes will be outside the membrane in the cytoplasm or in the core of the protein. Hydrophobic amino acids will be located in the bilayer. 4) IONIC INTERACTIONS - charge-charge interactions between a positively charged amino acid (ex: lysine) and a negatively charged amino acid (aspartic acid) (NH3+ R group interacting with COO- R group - not the zwitterion part) 5) VAN DER WAALS FORCES - intermolecular forces that repel atoms away from each other (steric hindrance) 6) PROLINE TURNS - proline's unusual shape will cause a KINK in the middle of an alpha helix, but it aids in beta turns (see proline) Quaternary Protein Structure - association of multiple folded proteins into a multi-subunit complex: Ex: Hemoglobin Hemoglobin - a layer of water that surrounds a dissolved protein. The water molecules in this layer interact closely with each other and with the protein's surface. The water in the hydration layer is more ordered than the bulk water in the general area and is considered not to participate with the bulk (a.k.a. unstructured) water when considering colligative (relating to the binding together of molecules) properties Entropy and Protein folding (protein folding interactions) - It is true that entropy of a protein decreases when it is folded (more ordered). The surrounding also matters however (part of the system). In an unfolded protein, hydrophobic portions of the polypeptide are exposed to water molecules which order themselves into hydration spheres (ordered). When a protein is folded, these hydration spheres break down (entropy of water molecules/surrounding increases). In other words, while entropy of the protein decreases (more stable) due to folding, the entropy of the surrounding region of water increases, leading to an OVERALL increase in entropy. Nonpolar solvation (very ordered water molecules) --> Polar/charged solvation (less ordered water molecules). Favorable INCREASE IN ENTROPY is a major CONTRIBUTOR to the OVERALL CONFORMATIONAL STABILITY of the folded protein. Increase in entropy is proportional to the conformational stability of a protein. Protein Denaturing Agents - -acid-pH (destroys tertiary and quaternary structure) -Chemicals (destroys secondary, tertiary, and quaternary structures) -Temperature (destroys secondary, tertiary, and quaternary structures) -Enzymes (destroys primary, secondary, tertiary, and quaternary structures) Protein separation technique: Isoelectric point (pI) - Isoelectric focusing is separating proteins by isoelectric point (pI). Proteins are placed in a gel with a stable pH gradient. If a protein is in a region of the gel with a lower pH than its pI, then the protein will be POSITIVELY charged and move towards the negative cathode. If a protein has a pH higher than its pI, it will be NEGATIVELY charged and move towards the positive anode. As the proteins move from their pHs to their respective pIs, their charges NEUTRALIZE and the protein will eventually stop to move (no charge, so no more pull towards either direction). Protein separation technique: Electrophoresis - Electrophoresis separates proteins based on size. Proteins are first placed in a detergent which DENATURES them and coats them with a NEGATIVE charge. This gives proteins a UNIFORM charge/mass ratio. Proteins are then run through a gel of polyacrylamide which slows down the migration of large proteins over the smaller ones. Gel is run from - to + electrodes and so the proteins are pulled towards the + electrode. The smallest proteins will run furthest through the gel, while the largest proteins will stay near the top. Central Dogma - DNA --> RNA --> Proteins DNA and RNA has NUCLEIC ACID building blocks Proteins have AMINO ACID building blocks Common Binding Proteins - Hemoglobin, Calmodulin, Troponin, Tropomyosin, Histones, Transcription factors, Cell adhesion molecules Immune System Proteins - Antigens (foreign bodies), Antibodies (produced in the body to fight off antigens) Structural Proteins - Actin (thin filaments, microfilaments), Tubulin (microtubules), Keratin (hair and nails, intermediate filaments), Elastin (connective tissue, extracellular matrix) Motor Proteins - Myosin (power stroke, cellular transport), Kinesis and Dyneins (vesicles, cellular transport, cell division, cilia, flagella) Kinesins - Move along microtubules from (CENTER OF CELL to PERIPHERY; NERVE CELL BODY --> DENDRITE) Dyneins - Move alone microtubules (+) to (-) end (PERIPHERY to CENTER OF THE CELL; nerve cell DENDRITE --> CELL BODY) Think Dyenin so D for dendrite Enzymes - PROTEINS increase reaction rate, lower activation energy, catalyze both forward and reverse reactions, NOT consumed in the process, NEVER alter thermodynamic properties of a reaction, and MOST NOTABLE DIFFERENCE: As proteins, enzymes are far more sensitive than inorganic catalysts to environmental conditions such as temperature and pH. Oxidoreductases (enzyme classification by reaction type): - REDOX reactions Transferases - Transfer of a functional group (e.g. kinases, aminotransferases) Hydrolases - Hydrolysis (cleavage with H2O) Isomerases - Rearrangements (e.g. phosphoglucose isomerase [G6P-->F6P], epimerases) Lyases - AB <--> A + B [cleavage/synthesis; No H2O, not hydrolysis] Stored in the liver and adipose tissue. Can lead to hypervitaminosis (toxic levels of vitamins) Antioxidants (vitamin E) Bloodclotting/blood pressure (vitamin K) Eyesight (Vitamin A) Calcium Levels (Vitamin D) Water Soluble Vitamins - All of the rest Water soluble vitamins act as cofactors Vitamins - small, ORGANIC molecules, essential nutrients necessary for metabolism. Humans have lost the ability to synthesize vitamins in sufficient quantities. We need vitamins in trace amounts so it takes less energy to obtain from DIET than to synthesize on our own. Vitamins have to be modified later in the body. Minerals - small, INORGANIC molecules, necessary for bone formation (calcium and phosphate), ion gradients (sodium and potassium), oxygen transport (iron containing heme group), muscle contraction (calcium), ATP processing (magnesium), stomach acid production (chlorine), etc. Obtained via DIET. Needed in small quantities (MICRONUTRIENTS) Steady State assumpuption Kinetics Formula: - E + S <--> ES --> E + P Michaelis Menten Equation - V = (Vmax*[S])/(Km+[S]) The relationship between REACTION VELOCITY, Km, and SUBSTRATE CONCENTRATION Vmax - the theoretical maximum rate of a reaction where an enzyme and substrate bind. At low [S], reaction rate increases rapidly because there is a higher chance for the free substrates to encounter an empty enzyme. As [S] increases, , the likelihood of encountering empty enzymes by free substrates decreases. At Vmax, all enzymes are bound to (preoccupied with) a substrate and as soon as a substrate is converted into a product, another substrate binds to the enzyme. Saturated - When all of the enzymes are preoccupied with substrates, the reaction is deemed SATURATED Michaelis Constant: (Km) - Km IS THE RELATIVE MEASURE OF AN ENZYME'S AFFINITY FOR ITS SUBSTRATED (Km = (k-1 + k2)/k1 Km = [S] @ Vmax/2 Lineweaver-Burk Plots - a double-interse graph of the reaction rate (v inverted to 1/v) and substrate concentration ([S] inverted to 1/[S]) Applications: Used to determine what kind of enzyme inhibition is occurring as we can easily calculate the IMPACT of an INHIBITOR on the Km and Vmax. Reversible Enzyme Inhibition - Inhibitor is not permanently bound to the enzyme; enzyme is not completely disabled Competitive Inhibition - INHIBITOR BINDS AT THE ACTIVE SITE. The inhibitor resembles the substrate in shape; the inhibitory effect CAN BE OVERCOME by INCREASING the CONCENTRATION OF SUBSTRATE Vmax = No Change Km = Increases Ex: Statin is a competitive inhibitor which binds to the active site of the HMG-CoA reductase enzyme and inhibits the enzyme's function of producing cholesterol in the liver. Chelators bind to the active site in order to be efficient Uncompetitive Inhibition - Inhibitor binds ONLY WITH THE ENZYME-SUBSTRATE COMPLEX. Typically RARE. Binds to another site which opens up when the ES complex is formed. Vmax = Decreases Km = Decreases Ex: Lithium is a drug used to treat maniac depression. Lithium acts as an uncompetitive inhibitor in the phosphoinositide pathway, inhibiting inositol monophosphate and preventing the inositol recycling in the brain. (inositol recycling is probably what is leading to the maniac depression) Non-Competitive Inhibition - INHIBITOR BINDS AWAY FROM THE ACTIVE SITE and changes the shape of the enzyme. The inhibitor has an EQUAL affinity for both the enzyme-substrate complex (E-S) and the enzyme (E). Vmax = Decreases Km = No Change Ex: Alanine acts as a non-competitive inhibitor for the enzyme pyruvate kinase. Pyruvate kinase normally transfers one phosphate group from PEP to ADP, creating pyruvate and one ATP. Alanine can bind to pyruvate kinase (whether or not PEP) the substrate is bound to inhibit activity. This creates a negative feedback loop since alanine syntheiss is derived from pyruvate. Mixed Inhibition - Ribose vs. deoxyribose - Glucose - Alpha glucose vs. beta glucose (ANOMERS) Animals have ALPHA glucose (OH on 1 C down) Plants have BETA glucose (OH on 1 C up) monosaccharide -OH groups on glucose can function as NUCLEOPHILES Fructose - Linear form is a ketose monosaccharide Galactose - monosaccharide Mannose - monosaccharide Lactose - Galactose + Glucose (beta linked by 1,4-glycosidic linkage) disaccharide Maltose - Glucose + Glucose Think single malt and double malt disaccharide Sucrose - Glucose + Fructose Think of coke in US (uses fructose) vs. coke in mexico (uses sucrose) disaccharide Pyranose vs. Furanose - Pyranose (6 member ring) Furanose (5 member ring). THINK F for Five Hemiacetals vs. Hemiketals - Hemiacetal (cyclical aldehydes) Hemiketals (cylical ketones) R/S vs D/L - NOT THE SAME THING! When focusing on R or S, rank the substituents by priority group (atomic number I think) When focusing on D or L, DRAW a FISCHER PROJECTION! If the hydroxyl group attached to the highest numbered chiral carbon is on the right in the Fischer projection, it is D-; if it is on the left it is L-. D-Sugars - ALL HUMAN BODY SUGARS are D-Sugars L sugars DO NOT OCCUR naturally in humans D-Glucose vs. L-Glucose - ENANTIOMERS (same molecule, but different stereochemistry) L - furthest -OH group from the CARBONYL is on the LEFT in the Fischer projection D- furthest -OH group from the CARBONYL is on the RIGHT in the Fischer projection Same molecule; different stereochemistry at the last chiral carbon Anomers - SAME molecule; different stereochemistry at the anomeric carbon Anomeric carbon = carbonyl carbon in the Fischer projection Epimers - Ex: Glucose vs. Galactose, Mannose vs. Glucose (Fischer projection) DIFFERENT MOLECULES, an example of diastereomers (R vs S configurations are different at one specific chiral carbon) NOTICE that ENANTIOMERS and ANOMERS have the SAME name (glucose in all of these examples), whereas the EPIMERS have DIFFERENT names (glucose/galactose) AMPHIPATHIC (carboxylic acid is polar) Surfactant - An amphipathic molecule secreted by cells in the alveoli (type II alveolar cells) that reduces surface tension on the inside of the alveolar walls. This prevents the alveoli from collapsing upon exhale and sticking together, thus reducing the effort required for inspiration. Similar to a fatty acid in structure (polar head and non polar tail) Glycerol - Triacylglycerol (Triglycerides) - Glycerol backbone with 3 fatty acids attached via ESTER linkages NOT Amphipathic Saturated vs. Unsaturated fats - Saturated fats are saturated with Hydrogen. Saturated fats do not have any double bonds. Unsaturated fats have double bonds (kinks) which can be in either the cis or trans orientation. Unsaturated fat is healthier since it generates fewer calories. Saturated fat has the most stored energy so it is great to consume under famine conditions. Trans-unsaturated fat cannot be broken down in the body and it accumulates in the blood. Trans-unsaturated fat is not created in the body and is ingested via processed foods. Your body can however breakdown ONLY the cis-unsaturated fat. Saturated fat is SOLID at room temperature as they typically have a HIGHER MELTING POINT. Saturated fat can lead to cardiovascular disease. Phospholipids - Two fatty acid moieties with a phosphate group that is attached to some other functional group (ex: like choline to make phosphatidycholine). The two fatty acid moieties and the phosphate group are attached to a glycerol backbone. HAS ESTER GROUPS AMPHIPATHIC (phospho group is polar) Saponification - The hydrolysis of an ester (base promoted ester hydrolysis used NaOH or something like that). Triglycerides, phospholipids, etc can be hydrolyzed Steroids - Tetracyclic (4) ringed. NOT Amphipathic Terpenes - Polymer of ISOPRENES (can be a chain or a ring) A terpene is a repetitive chain of hydrocarbons (usually multiples of 5) Terpenoids - Terpene with OXYGEN NOT Amphiapthic Sphingolipids - sphingomyelins (phosphate group on 4th carbon on the backbone is attached to choline or ethanolamine, make up the myelin sheath. It's breakdown results multiple sclerosis) or cerebrosides (4th carbon on the backbone is attached to a sugar monomer) AMPHIPATHIC (oxygen and nitrogen are polar) Waxes - Waxes consist of a longchain fatty acid linked through an ester oxygen to a long-chain alcohol. These molecules are completely water-insoluble and generally solid at biological temperatures. Their hydrophobic nature allows them to function as water repellents on leaves of some plants, on feathers, and on the cuticles of certain insects. NOT Amphiathic Glycolipids - Phosphatid (glycerol backbone that has 2 fatty acid moieties and phosphate group) where the phosphate group is attached to a sugar monomer Prostaglandins (PGX) - Play a key role in inflammation, blood pressure, blood clotting, fever (good when fighting infection), pain, labor, sleep-wake cycle. NOT Amphipathic Lipid mediators that have AUTOCRINE (self-target) and PARACRINE (target = cell in immediate vicinity) functions throughout the body UNLIKE ENDOCRINE HORMONES: -Produced and released throughout the body;NOT only in specialized glands -Act LOCALLY, rather than traveling to a distant target via the bloodstream. Molality - (moles of solute)/(kg. of solvent) Less concentrated than molarity