MCB 2000 Exam 1

MCB 2000 Exam 1 Alanine (ala) nonpolar aliphatic Glycine (gly) nonpolar aliphatic Valine (val) nonpolar aliphatic branched chain Leucine (leu) nonpolar aliphatic branched chain Isoleucine (ile) nonpolar aliphatic branched chain Proline (pro) nonpolar aliphatic ring structure that is not aromatic Phenylalanine (phe) nonpolar aromatic permits absorption of UV light at 280 nm Tyrosine (tyr) polar and nonpolar properties -amphipathic uncharged aromatic OH group Tryptophan (trp) nonpolar aromatic permits absorption of UV light at 280 nm Serine (ser) polar uncharged OH group Threonine (thr) polar uncharge OH group Cysteine (cys) polar uncharged SH group -groups can become oxidized and form disulfide bonds to each other (S-S) Asparagine (asn) polar uncharged carboxamide group Methionine (met) nonpolar aliphatic thioether Glutamine (gln) polar uncharged carboxamide group Lysine (lys) polar charged positive Arginine (arg) polar charged positive Histidine (his) polar charged positive pKa 6, close to physiological can act as acid or base Aspartic acid (asp) polar charged negative Glutamic acid (glu) polar charged negative What makes water essential for life? -solvent -organizing principle -can be reactant or product in reaciton Common electronegative atoms in biological systems OxygenNitrogenSulfur Types of Noncovalent Interactions Hydrogen Bonding Electrostatic, Ionic interactions Van der Waals force Hyrdrophobic interactions Hydrogen bonds btwn electronegative atom (acceptor) and an H atom that is covalently bonded to another atom (donor) -dont last very long -weak association -bond is strongest at 180 degrees Importance of Hydrogen bonds -solvent properties of water -structure of proteins and nucleic acids Electrostatic, Ionic interactions Occur btwn atoms with a complete - charge and complete + charge -strength of interaction determine by distance and solvent Importance of electrostatic interactions protein-protein interactions DNA protein interactions Catalytic mechanisms Van der Waals forces charge fluctuations transiently produces + or - poles even in nonpolar molecules -weakest -present with snug fit btwn atoms -interactions in membranes - contributes to how permeable it is Hydrophobic interactions Nonpolar molecules that can not form H bonds -form spontaneously Hydrophobic interactions importance allow for an increase in entropy in water The hydrophobic effect Entropy driven association -structure function of proteins - hydrophobic amino acids on inside Importance of noncovalent interactions flexibility stability to large molecules essential to specificity 4 properties of all living organisms Chemical and energy transformation adaptation replication organization and complexity What determines polarity electronegative atoms -electrons not being shared equally Properties of polar molecules partial charges noncovalent interactions hydrophilic reactivity amphipathic both polar and nonpolar molecule Myoglobin stores O2 and releases it under extreme excursion globular protein that is hydrophobic on inside has 8 alpha helixes has high affinity to O2 bc it is tertiary and cant do coopertivity Reversible ionization unique property of water H2O - H+ + OH- Molarity of water 55.5 M Keq of water 1.8 x 10^-16 -indicates that number of dissociate molecules are very small -always products over reactants Kw 1 x 10^-14 will always be the concentration of H+ times concentration of OH- -central role of water as biological solvent pH -log[H+] range 0-14 How does Metabolism change pH? Metabolism produces changes in [H+] which would make the body very acidic without buffers Changes of pH (by .2 -.3) in body Is a signaling mechanism Promotes cell proliferation, migration, assembly of actin Regulates specific proteins binding affinity, activity, recruitment High pH can cause what disease? Cancer Low pH can cause what disease? Neurodegenerative diseases Metabolic acidosis acid-base pathological disorder where there is an increase in the production of acid or a loss of bicarbonate -may cause lactic acidosis, ketoacidosis, complication with drugs, diarrhea, and altered kidney function Acid substance that can release H ion Base substance that can accept H ions Ka expression of the extent of ionization dissociates on top, acid on bottom -used to compare strengths of acids Henderson-Hasselbalch equation pH = pKa + log [A-]/[HA] Applies to weak acids and bases Indicates relationship btwn Ka and pH When pH = pKa the acid is 50% dissociated will be the point of inflection on a concentration curve How is knowledge of pKa important in aspirin? It's a weak acid with pKa 3.5 and has hydrophobic character. Absorbed through cell lining the stomach and small intestine Absorption requires passage through plasma membrane which rate is determined by polarity and charge of molecule. When aspirin is protonated it can pass through membrane since it has no charge. It will be absorbed better through stomach since it has a low pH. Selective avantage for cancer cells proliferation avoiding apoptosis migration and invasion avoiding immune detection Intra and extracellular pH of normal cells IC pH ~ 7.2 EC pH ~ 7.4 Intra and extracellular pH of cancer cells IC pH 7.4 EC pH ~ 6.7-7.1 Will have a reverse pH gradient that benefits the cells Equilibrium no net change in concentration Buffer that your body uses Bicarbonate Amino group weak organic base Carboxylic group weak organic acid Which carboxylic acid will lose a proton first in glutamic acid? the one closer to the amino group will lose it first bc the influence of the base will cause it to lose it to the immediate environment Point of best buffer capacity when pH = pKa What type of metabolism do cancer cells use? Only glycolysis, even when oxygen is present. This requires a lot of energy, since only 2 ATP are created per glucose. pyruvate is then exported in the form of lactic acid from the cell Why reverse gradient benefits cancer cells Immune cells do not like acid environment so they wont attach the cells Doxorubicin chemotherapy agent that is hydrophobic with rings but has a lot of polar groups its absorption has to do with amine group pKa 8.2 Will pick up a proton in acid environment of cancer cell and be less likely to enter the cell than normal cell Groups of cells that doxorubicin will affect most Cells that divide rapidly -blood, hair, intestines Chemo drugs with basic vs acidic functional groups Basic functional groups are more likely to pick up a proton in the extracellular environment of cancer cells, therefore not being able to enter the cell. Acidic functional groups will not have this problem Homeostasis to stay around a certain set point Buffer mixture of undissociated acid and its conjugate base Resists changes in pH when either H+ or OH- is added Phosphate buffer used in intracellular fluids to keep pH 7.2 Carbonic acid/ bicarbonate buffer in blood Works bc of high CO2 dissolved in body fluids and equilibrium btwn dissolved CO2 and CO2 in lungs How buffers function involves 2 reversible equilibria, one involving water What factors influence the effectiveness of a buffer Concentration pKa close to wanted pH Essential Amino acids Histidine Isoleucine Leucin Lysine Methionine Phenylalanine Threonine Tryptophan Valine -cannot be made in body, must come in diet Amino acids Are ionizable Can be a buffer Nonessential amino acids Can be created through TCA cycle intermediates Structure of amino acid at pH 7 Deprotonated carboxyl group and protonated amino group (Zwitterionic form) -no net charge pKa of carboxyl group in amino acid between 2-3 Will be protonated below this, and depro above pKa of amino group in amino acid between 9-10 will be protonated below 9 and depro above Ionizable R groups of amino acids The amino acid will have a third pKa for their side chain which will contribute to noncovalent electrostatic interactions in proteins Amino acids with ionizable side chains Arginine Lysine Tyrosine Cysteine (Terminal alpha-amino group) - NH3+ Histidine Aspartic acid (aspargine) Glutamic acid (glutamine) (Terminal alpha-carboxyl group) Imidazole group 5 membered ring of histidine can bind or release protons at neutral pH Can act as acid or base -important in protein structure and function At pH pKa H+ on, protonated At pH pKa H+ off, deprotonated Optical activity of amino acids They have two chiral forms, L and D. With the H in the back, L is when amine group is on left Only L isomers found in proteins aliphatic side chain of carbons arranged in linear or branched chain structure Aromatic side chain of carbons arranged in a resonance stabilized ring structure with conjugative bonds Can absorb ultraviolet light in 280 nm bc of resonance structure Amino acids with branched structures important for skeletal muscle bc it can make energy by breaking them down found abundant in muscle valine leucine isoleucine Peptide bonds covalent link btwn amino acid chains where carboxylic acid of one gorup and amino group of other bond Is an amide bond Direction of peptide chain formation N-C Proteins emerge from ribosome amino group first N end will have + charge, C will have - charge making the chain net charge 0 without R group consideration Formation of peptide bond A condensation reaction where water is eliminated. Carried out by catalytic RNA to ensure bond stays since reactants are favored Catalytic RNA found in ribosome, are used to put energy into peptide bond formation Use 6 ATP/GTP to form one bond Characteristics of peptide bond It is planar has two resonance states (shown) and will constantly flip btwn these states creating a partial double bond character so it will remain rigid and no rotation will occur - Where does rotation occur around a peptide bond phi angle of C-N and psi angle of C-C -allows interaction of the side chains of amino acids -will create cis and trans structures, but cis isnt common bc of steric effects Ramachandran plot Representation of permissible psi and phi angles at which a protein can fold and be stable without steric effects Steric effects when the components of a molecule are in close proximity to each other resulting in strain and instability of the molecule When does protein start folding? As it exits the ribosome Ribosome covers 30 amino acids, so when first 30 come out folding begins As folding goes on enzymes will tweak the structures Diseases of protein misfolding Parkinson's and Alzheimers Form larger fibrous structures Cell attempts to get rid of misfolded proteins but in this case they can't Primary protein structure Linear sequence of amino acids Secondary protein structure Involves hydrogen bonding along polypeptide backbone btwn NH and CO, no R group interaction Alpha helix, beta sheets, and beta turns Tertiary protein structure Involves noncovalent interactions btwn amino acid R groups and the pattern of disulfide bonds Amino acids far apart in primary structure are brought closer and interact via noncovalent interactions Nonpolar amino acids will go toward center Quaternary protein structure 2 or more polypeptide chains (subunits) stabilized by noncovalent interactions and sometimes disulfide bonds Interact to form the functional protein Alpha helix COO- H bonds with NH4 at 4 pairs away The O binds to H Repetitive H bonding creates a helix R groups project outward Constraints on alpha helix Steric effects cause valine isoleucine threonine to be less common Reactivity effects causes some side chains that are H bond donors or acceptors to compete with main CO and NH groups (serine, aspartic acid, asparagine) Psi angle in proline not consistent with helix Glycine destabilizes H side chain bc it is to small Beta strand/sheet multiple beta strands make beta sheets R groups will project above and below chain to accommodate for large ones Have directionality H bonds are perpendicular to polypeptide chain Antiparallel beta sheet H bonds are perpendicular Parallel Beta sheet H bonds are slanted Ribbon diagram shows directionality of beta sheets when connected by H bonds Domains unique structural entity that may have formed through cut and paste Used to identify part of protein that has different folding pattern, function, etc In tertiary structure ATP binding domain part of a protein that binds to ATP p53 beta barrel that binds to DNA mutation may cause tumors loop 2 position it on DNA Anfinsen's Experiment Used Ribonuclease (RNase) to determine the factors of how a protein folds done in vitro Significance of ribonuclease in experiment it is small and its activity can be measured bc it is an enzyme that degrades RNA -120 amino acid chain with 4 disulfide bonds How disulfide bonds form oxidation reaction Lose H's and S's form a covalent bond How to break disulfide bonds through a reduction reaction using a strong reducing agent Disulfide bonding in vivo Done in the lumen of the ER where there are enzymes that make sure the correct cysteines pair together correctly carried out non spontaneously Anfinsen's experiment - two reagents 8M urea beta - mercaptoethanol 8M urea used in Anfinsen's experiment to cause ribonucleases H bonds and electrostatic interactions to unfold Beta-Mercaptoethanol reagent used in excess in Anfinsen's experiment to cause ribonucleases disulfide bonds to break apart Dialysis bag bag with small pores which small molecules can escape Conclusions from Anfinsen's experiment Protein folding patterns are dependent on the amino acid sequence -folding process will begin before disulfide bond formation When partially active ribonuclease is treated with trace amounts of beta-merc the proteins that are folded correctly will stay in state bc they are stable. Proteins folded wrong will unfold for a chance to refold correctly Can Anfinsens experiment be done with heat instead of reagents? For this protein yes, however some proteins are irreversible when denatured with heat If only one domain in a protein folds correctly, how does it refold? the domain that is correct will remain folded, and the rest will unfold and refold Protein Denaturation lose all structure of protein except for primary peptide sequence Protein Misfolding protein will still have structure but it may be wrong or missing something Things a cell does with misfolded proteins refolds targets for degradation Chaperonins large bullet shaped protein complexes that help fold proteins that cant fold on one increase rate of protein folding -increase in numbers when exposed to heat shock to deal with high temp effects -accumulate in nucleolus to protect RNA and other proteins Proteasomes Large protein complex that degrade proteins into free amino acids Reason sickle cell point mutation causes disease Glutamic acid is a polar negative amino acid on the surface of hemoglobin and is replaced by a nonpolar valine which will try and stick to other nonpolar valines and cause polymers of beta chains Non conserved mutation replacing a very different amino acid for the one that should be there Amyloid segments short sequences of 6 amino acids that are typically inside a sequence of a protein but if they are exposed to the surface they become sticky to other segments and form rope like structures of beta strands which extensive H bonding that is very hard to degrade Psi angle between the carboxyl and the Alpha carbon Phi angle between the nitrogen and the alpha Carbon Type of amino acid you would find in membrane bound protein Nonpolar amino acid bc it is hydrophobic and will hide in lipid bilayer First proteins in which structural studies were done hemoglobin myoglobin Crystallography proteins are crystallized to determine structure Similarities btwn hemoglobin and myoglobin evolutionarily related amino acid sequence similar throughout have heme groups Why does hemoglobin contribute to blood buffer? It can transport CO2 which is part of the bicarbonate buffer Alpha and beta subunits of hemoglobin about the same size as myoglobin individually encoded by two separate genes -must be equal amounts encoded Thalassemia disease where to much of one chain of hemoglobin is produced will be hemoglobins with one type of chain missing Why is a heme used for oxygen binding? oxygen is poorly soluble in water, and would not make it to far target tissues Metals have strong tendency to bind to oxygen but are very reactive Iron is sequestered in heme to make it less reactive Structure of heme planar and hydrophobic Iron surrounded by 4 nitrogens which are electronegative and will keep iron in Fe2+ state which binds O2 Why does heme have reversible O2 binding? it is sequestered within proteins structure which will prevent full transfer of electrons when binding to O2 Molecular structure of heme when in deoxy state Iron hangs below plane of ring, still bound to 4 N's Imidazole group of histidine hangs below (proximal histidine) distal histidine is on other side but not bonded has three noncovalent interactions to stabilize this state Three noncovalent interactions in deoxy heme Aspartic acid electrostatic interaction with histidine (salt bridge) Salt bridge btwn histidine and lysine valine carbonyl group forms H bond with tyrosine Molecular structure of heme when in oxy state Fe is in plane of ring, dragging histidine and breaking noncovalent interactions structure of Hb will rotate 15 degrees -central cavity gets smaller CO binding to hemoglobin has 200x greater affinity than O2 -Will cause Hb to not release other O2 symptoms are cold/flu like Will cause poisoning -wont occur in low levels of CO bc distal histidine creates steric effects to reduce probability of it binding Allosteric behavior Changing to other shapes -protein can only do this if its quarternary -protein will have sigmoidal binding curve Cooperativity when O2 binds to Hb on one subunit it is relayed to other subunits Sigmoidal curve S shaped binding curve -hemoglobin curve Hyperbolic curve binding curve that rises quickly and plateaus at top -myoglobin curve Partial pressure in tissues 25 When will myoglobin release oxygen? When partial pressure goes below physiological levels -look at saturation curve T state of Hb deoxy state R state of Hb oxygen bound HB Which state of Hb does CO stabilize? the R state -therefor oxygen will stay bound to Hb Which state of Hb does 2-3biphosphoglycerate stabilize? T state will allow Hb to release O2 What type of interaction does heme have with hemoglobin? hydrophobic interaction pH optima determined by what interactions? electrostatic 2-3Biphosphoglycerate (23BPG) binds to central cavity of Hb when oxygen is released and stabilizes T state Major factor in releasing oxygen Interacts electronegatively with positive charges of histidine and lysine since it is negative 23BPG in fetal hemoglobin fetal hemoglobin has a serine instead of histidine, causing less interaction with 23BPG and therefore keeping it in the R state Physiological relevance of fetal hemoglobin fetus must have higher affinity for oxygen bc it gets oxygen from maternal circulation by capturing it 23BPG is produced where? as an intermediate in glycolysis and has a 1:1 ratio with hemoglobin What type of metabolism to RBC use? Glycolysis only Bohr effect regulation of oxygen binding to Hb by H+ and CO2 which are biproducts of metabolism Carbonic anhydrase enzyme in RBC that increases the rate of conversion of CO2 and H2O to H2CO3 Decrease in pH does what to oxygen saturation curve? Shifts it to the right favors T state Increase in CO2 in blood does what to oxygen saturation curve? Shifts it to right favors T state Caused by Bohr effect because CO2 will have greater affinity to Hb than O2, causing oxygen release Allosteric effectors of Hb CO2, H+, and 23BPG will all cause right shift in graph and favored T state Acid base property of histidine in hemoglobin Histidine will pick up a proton in the tissues (acting as base) and release its proton in lungs (acting as acid) When histidine has its proton the salt bridge btwn aspartic acid is present. It breaks when oxygen binds and proton is released Why do changes in pH affect the affinity of Hb for hemoglobin? Bc when a proton binds to histidine, oxygen will be released, meaning the T state will be stabilized Allosteric effects of CO2 binds to the amino terminus of each Hb chain and converts it into a carbamate which will give it a charge and release proton which will interact to establish a salt bridge Why is blood buffer's pKa value 6.35 when bicarbonate typically has pKa around 3? there is a lot of CO2 dissolved in the blood and not as much HCO3- causing pKa to be higher A sigmoidal curve indicates what? cooperativity Prion protein protein found in central nervous system that normally has a alpha helical structure and one beta sheet. A disease confirmation will have multiple beta sheets and can enter other cells to influence other proteins to switch to disease confirmation through amyloid formation Alpha-synuclein another protein that has prion like behavior and contributes to parkinson's disease Three possible mechanisms of reducing amyloids Enzymes that specifically disaggregate amyloids Use small molecules as extra chaperones to lower probability the proteins will be exposed to amyloidogenic segments A compound to cap steric zippers of amyloid fibers slowing their formation