BSC 450 Final Exam Study Guide
BSC 450 Final Exam Study Guide BSC 450
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This 23 page Study Guide was uploaded by Jordana Baraad on Monday December 7, 2015. The Study Guide belongs to BSC 450 at University of Alabama - Tuscaloosa taught by Dr. Ramonell in Summer 2015. Since its upload, it has received 129 views. For similar materials see Fundamentals of Biochemistry in Biological Sciences at University of Alabama - Tuscaloosa.
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Study Guide for Chapter 14: Glycolysis – BSC 450/550 1. Describe and understand the importance of phosphorylated intermediates in glycolysis. a. All 9 glycolytic intermediates are phosphorylated; serve 3 functions i. Addition of phosphorylation prevents sugar from leaving the cell 1. Plasma membrane lacks transporters for phosphates sugars 2. No further energy expended to retain sugar intermediates 3. Ex. G6P ii. Enzymatic conservation of metabolic energy 1. Energy released in breakage of phosphoanhydride bonds (e.x. hydrolysis of ATP energy release) 2. Partial conservation in formation of phosphate esters (i.e. G6P) iii. Phosphate groups + enzyme active sites binding energy lowered activation energy & increased reaction specificity (incorporation of Mg + essential to function) b. 6 PARTICULARLY important intermediates & their functions: i. Glucose6phosphate (G6P): keeps glucose from leaving the cell; maintains concentration gradient so rxn can procede ii. Fructose1,6Bisphosphate: ONLY molecule that can go into pyruvate production pathway st 1. 1 committed step of glycolysis iii. Glyceraldehyde3Phosphate (G3P): oxidized to form NADH iv. 1,3BPG: key intermediatedephosphorylated to produce 2 ATP v. Phosphoglycerate (PGA): highenergy molecule, 2 ATP produced 1. Catalyzes subsequent glycolysis step vi. PEP: highenergy molecule, 2 ATP produced from dephosphorylation pyruvate 2. Know the enzymes involved in glycolysis. Know their associated cofactors, what reaction they perform and note any unique aspects of the reaction mechanism. a. ***Hexokinase: phosphorylation of glucose @ C6 Glucose6phosphate(G6P) 2+ i. Coordinated by Mg b. Phosphoglucose isomerase: G6P F6P i. Moving carbonyl and hydroxyl grops 1. Forms linear enediol intermediate 2. Avoid negative buildup 3. Step 1: binding/opening ring a. His’s in active site coordinate ringopening b. Activesite base glutamate w/n enzyme 4. Step 2: proton abstraction—forms intermediate c. *** Phosphofructokinase1 (PFK1): F6P Fructose1,6Bisphosphate (F1,6 BP) i. TIGHTLY regulated enzyme ii. Mg as cofactor == stabilizes ATP 1. Neutralization interactions w/ alpha and beta phosphates 2. Acts like “giant proton” d. Aldolase: F1,6BP cleaved into 2 triose molecules (G3P & DHAP) i. (not favorable) ii. 2 classes: 1. plants/animals: enolate & Schiff base intermediate a. covalent catalysis w/ Lys @ active site 2. fungi/bacteria: enolate intermediate a. metal ion assisted catalysis w/ Zn io in active site to coordinate w/ carbonyl and stabilize enolate e. Triose phosphate isomerase: interconvert triose phosphates f. Glyceraldehyde 3phosphate dehydrogenase: G3P 1,3BPG i. Carbonyl hydroxyl functional gropu on C1 g. Phosphoglycerate kinase: 1,3BPG 3 Phosphoglycerate (PGA) + NADH i. Step 1: enzymesubstrate complex formation 1. Activesite Cys 2. More reactive form: reduced pH, NAD+ bound, thiolate form ii. Step 2: covalent thiohemiacetal linkage btwn substrate & S in Cys iii. Step 3: enzymesubstrate complex oxidized by activesite NAD+ iv. Step 4: NADH leaves; immediately replaced v. Step 5: phosphorolysis of covalent thioester linkage release of of 1,3 BPG h. Phosophoglycerate mutase: 3PGA 2PGA i. Isomerization rxn involving transfer of functional group ii. 2 His activesite residues 1. 1 part of –OH transfer C2C3, other is base then acid catalyst i. Enolase: 2PGA phosphoenol pyruvate (PEP) i. Dehydration rxn j. ***Pyruvate kinase: dephosphoryl2+ion+PEP Pyruvate i. Metalassisted rxn: Mg , K , Mn coordinate charges in active site to add phosphate to ADP k. “***” indicates tightly regulated enzyme 3. Be able to apply common reaction mechanisms to new metabolic problems. Be able to describe what type of reaction or mechanism would be required to reach a given molecule. a. Common reaction mechanisms: i. Phosphorylation/ dephosphorylation: done w/ kinase (either adds P to i sugar OR removes from sugar to add to ADP ATP 1. Indicative of substratelevel phosphorylation ii. Oxidation: occurs via dehydration by enzyme “dehydrogenase” + 1. Often done to substrate so NAD can be reduced NADH and act as electron carrier iii. Isomerization: generally done w/ “isomerase” enzyme iv. Cleavage: enzyme ending w/ “ase” (ex. aldolase) 4. Know the four basic problems that must be addressed in glycolysis and how they are solved. a. Preparatory phase (Problems 1 &2) i. transform 6C molecule (glucose) 2 3C molecules (G3P & DHAP) ii. G3 + DHAP 2 G3P 1. Conversion to avoid wastefulness of 2 pathways pyruvate 2. Isomerization rxn b. Payoff phase (Problems 3& 4) i. Generate ATP for cell 1. Substratelevel phosphorylation 2. Must generate highenergy molecules: PEP + 1,3BPG ii. G3P pyruvate 1. Simplest steps possible to generate pyruvate + highenergy molecules 5. Understand how NAD and NADH work in the cell. + a. In glycolysis, NAD reduced NADH to be an electron carrier in CAC i. Reduction of NAD achieved by oxidation of sugar intermediate 1. Transfer of hydride ion ii. NADH =+reduced coenzyme iii. NAD = hydrogen acceptor bound to ROSSMAN FOLD b. NAD concentration in cell (< 10 M)much smaller than amount glucose metabolized in a few minutes i. Continuous reoxidation & recycling of NADH necessary for continuation of glycolysis 6. Understand how high energy compounds are generated in glycolysis and why they are formed. a. High energy compounds: PEP; 1,3BPG, pyruvate (final product of glycolysis, but still has lots of energy to be harvested—addressed in CAC) i. Formed by phosphorylation 1. Attack by P oi covalent thioester substrateenzyme complex b. Formation of highenergy products in glycolysis: exergonic; coupled to endergonic ATP production (ATP production could not occur, otherwise) c. Formed so that they can be dephosphorylated to phosphorylate ADP ATP i. Substratelevel phosphorylation—occurs twice in glycolysis 7. Understand the difference between substrate level phosphorylation and oxidative phosphorylation. a. Energy production i. Substratelevel: only net production of 2 ATP (4 produced – 4 required) 1. ATP produced by transfer of phosphoryl group from substrate (i.e. 1,3BPG) to ADP ii. Oxidative: LOTS produced b. Process/ location i. Substrate level: cytoplasm (phosphate transfers in glycolysis) ii. Oxidative: on cristae in mitochondria (electrochemical gradient) 8. Know the various fates of pyruvate in the cell. i. Hypoxic / anaerobic conditions fermentation ii. 2 Lactate (vigorously contracting muscles, erythrocytes, some microorganisms) OR 2 ethanol + 2 CO in y2 st iii. NAD reoxidized iv. Requires 15x more sugar for same energy output b. Aerobic conditions 2 AcetylCoA citric acid cycle 4 CO2 + 4 H20 Study Guide for Chapter 16: The Citric Acid Cycle – BSC 450/550 1. Understand the structure and functions of the enzymes in the pyruvate dehydrogenase complex (and αketoglutarate dehydrogenase complex) including their associated cofactors. a. E1: pyruvate dehydrogenase: decarboxylation rxn of pyruvate i. Decarboxylation due to thiazolium ring of TTP ii. PD transfers hydroxyethyl to E2 iii. Structure: 1domain enzyme, centrall bound TPP at active site iv. Cofactor: Thiamine pyrophosphate (TPP) 1. Electron sink, contains thyazolium ring b. E2: dihydroplipoyl transacetylase: reduce Acetyl CoA i. Transfers C2 group to CoA 2 Acetyl CoA ii. Acetyl CoA can enter Citric Acid Cycle) iii. Transfers some electrons to NAD energy 1. Yields 2 NADH iv. Structure: HUGE protein complex—60 subunits 1. 3 functionally distinct domains a. lipoyl domain: aminoterminal, containing lipoylLys residue b. E 1and E 3binding domain (central) c. Acyltransferase domain (core), contains acyltransferase active site v. Cofactors: CoA, lipoamide 1. Depends on cooperative binding to function a. Intermediates too reactive to bind, otherwise 2. Lipoyllysyl moiety: prosthetic group of dihydrolipoyl transacetylase a. Lipoate + Lys side chain = long, flexible arm that swings from active site E1 o active sites of E2 and E3 c. E3: dihydrolipoyl dehydrogenase i. Regenerates NADH immediately after it’s been reduced ii. Reduces E2 iii. Structure: 1domian protein; centrally bound FAD at active site iv. Cofactors: FAD, NADH 1. NADH carriers electrons produced to CAC 2. Oxidation of FAD resets E3 2. Understand the chemical roles and unique attributes of the enzymes involved in the Citric Acid Cycle. st a. Citrate synthase: catalyzes 1 rxn in CAC i. Rxn = Claissen condensation of AcetylCoA + Oxaloacetate citrate b. Aconitase: isomerizes tertiary hydroxyl group of citrate (2 step process) i. Step 1: dehydration 1. Citrate cisaconitate intermediate (source of enzyme name) a. H & OH removed ii. Step 2: rehydration 1. Intermediate isocitrate (H & OH added back differently) 2. Even w/ catalysis, not a favorable rxn a. Relies on low oxaloacetate concentration iii. Uses FeS cluster; regulates Fe homeostasis 1. Central Fe key to enzymestunction—balances negative charge c. Isocitrate dehydrogenase: catalyzes 1 decarboxylation of isocitrate i. Isocitrate alphaketogluterate (+ CO2 + NADH) ii. 3 steps 1. oxidation sets stage—hydride transfer to cofactor NAD(P)+ 2. decarboxylation: Mn = essential coordinating metal cofactor 3. rearrangement of enol d. Alphaketogluterate dehydrogenase complex: catalyzes 2 decarboxylation i. Alphaketogluterate succinyl dehydrogenate (HIGHLY reactive) ii. 3 enzymes (like in CAC entry rxn & photosynthesis, w/ same cofactors) 1. E1: alphaketogluterate dehydrogenase a. Coenzyme: TPP b. Decarboxylation rxn of alphaketogluterate 2. E2: lipoyl acetyl transferase a. Coenzyme: CoA, lipoamide / lipate (swinging arm) b. Add CoA 3. E3: Lipoyl dehydrogenase a. Coenzyme: FAD, NAD b. Generates NADH c. SuccinylCoA product continues in cycle e. SuccinylCoA synthetase: harvests energy from SuccinylCoA i. Energy in GTPform—immediately converted ATP ii. Enzyme intermediate = phosphohistidyl enzyme 1. His = essential residue iii. Generates succinate, which continues in cycle 1. Succinate production marks beginning of recycling stage f. Succinate dehydrogenase: oxidizes Succinate transfumarate i. FAD as cofactor—FADH2 produced ii. Stereospecific product g. Fumarase: hydrates fumarate Lmalate i. Stereospecific reactant and product—VERY specific, highly evolved h. Malate dehydrogenase: oxidizes Lmalate oxaloacetate i. Oxaloacetate regenerated—restart cycle w/ initial electron acceptor ii. NAD as cofactor—NADH produced 3. Understand the types of reactions that take place in the Citric Acid Cycle and know the final products of the cycle. a. Products: NADH, FADH , GTP2 (regeneration oxaloacetate), 4 CO , 4 H 2 2 b. Reaction types: i. Claissen condensation: citrate synthase ii. Dehydrationoxidation: aconitase & (fumarase + malate dehydrogenase) iii. Carboxylation: isocitrate dehydrogenase, alphaKG dehydrogenase complex iv. Oxidation: succinate dehydrogenase 4. Understand the role of Aconitase in iron homeostasis. a. Aconitase has FeS cluster in center: acts in formation of enzymesubstrate complex and in catalytic addition or removal H O2 b. Fedepletion in cells cells FeS cluster disassembly loss of aconitase activity c. Aconitase w/o FeS = apoaconitase—has new capability i. Binds to mRNA sequence encoding ferritin and the transferrin receptor, which effect iron mobilization and iron uptake ii. Iron regulartory proteins (IRP1 and IRP2) bind to these sequences (Iron response elements or IREs) to block ferritin synthesis iii. Apoaconitase binds to IREs instead of IRPs more efficient iron uptake and reduced iron storage in Fedeficient cells iv. Normalization of cellular Fe concentration IRP1 converte to aconitase (w/ FeS cluster) 1. IRP2 undergoes proteolytic degradation, ending lowFe response 5. Know the difference between a biochemical pathway and a biochemical cycle. a. Biochemical pathway: series of chemical reactions occurring within a cell i. Catalyzed by enzymes; each product becomes substrate for subsequent enzyme ii. Reactant DOES NOT = product(s) iii. Typically regulated by feedback inhibition iv. Ex. glycolysis b. Biochemical cycle: i. Also enzymecatalyzed w/ each product becoming substrate for subsequent enzyme—INCLUDING THE LAST ONE ii. End product of reaction is a reactant or substrate that starts the pathway iii. Regeneration built into cycle (not separate feedback inhibition process) iv. Ex. CAC 6. Understand the unique attributes of the enzyme fumarase. a. HIHGLY stereospecific enzyme—w/ respect to reactant and product i. Catalyzes hydration of the double bond of fumarate (but not the double bond of its cis isomer maleate) Lmalate ii. Rxn is reversible, but equally stereospecific 1. Fumarase will only act upon Lmalate; Dmalate not a substrate Study Guide for Chapter 19: Oxidative Phosphorylation 1. Understand the anatomy of the mitochondria and the organization of the membranes a. outer membrane: freely permeable to small molecules and ion i. porins: integral membrane proteins allow most ions below certain size in b. inner membrane: impermeable to most ions/ particles (even protons) i. require specific transport ii. contains: respiratory electron carriers (Complexes IIV), ADPATP translocase, ATP synthase (F F0 1specific transport for protons), other membrane transporters c. matrix: interior of mitochondria; site of CAC i. contains: pyruvate dehydrogenase complex, CAC enzymes, fatty acid beta oxidation enzymes, amino acid oxidation enzymes, DNA, ribosomes, ATP/ ADP, metal ions, many enzymes, soluble metabolic intermediates d. cristae: continuous w/ inner membrane houses thousands of copies of ETC (Oxidative phosphorylation) 2. Know the proteins and cofactors / prosthetic groups involved in oxidative phosphorylation th 4 complexes build proton gradient (though sometimes, a 5 is counted in with them) 1. NADH dehydrogenase –Complex I a. Cofactor: FMN (flavoprotein)—primary electron acceptor from NADH (1 e ) b. FeS Cluster (also present in Complexes IIIV) i. Single electron carriers ii. Coordinated by Cys in protiens iii. His Rieske FeS proteins iv. Same # Fe and S molecules v. Moving from matrix intermembrane space 1. Chain electrons accepted / donated to move across membrane 2. Succinate dehydrogenase – Complex II a. Cofactor: FAD (flavoprotein)—primary electron acceptor, becoming FADH 2 3. Cytochrome bc –1omplex III a. Cytochromes = hemecontaining proteins: Types a, b, c i. A,b,c differ by ring additions (Fecoordinated porphirin ring) ii. Hydrophobic isoprenoid tail for docking b. CoQ / Ubiquinone—CIII does Q cycle i. Semiquinone: single electron carrier 1. Hangs around to pick up 2n electron carrier 2. Picks up protons to balance charge 3. Docks elsewhere via hydrophobic tail 4. Cytochrome oxidase – Complex IV a. Also contains cytochromes (see above) 5. ATP Synthase – Complex V Part of respiratory chain, not ETC 3. Understand and be able to describe electron flow through the proteins / cofactors / molecules involved in oxidative phosphorylation. –Q CYCLE!!!! a. CI & CII Q (a.k.a. ubiquinone) i. pass fr NADH thru flavoprotein w/ cofactor FMN to FeS center N2 (CI) Q 1. electrons go thru C1 if derived from betaoxidation, succinate oxidation (CAC), cytosolic rxns ii. succinate binding site on subunits flavoprotein w/ cofactor FAD and series of FeS cluster on subunits A& B (CII) Q (bound to B) 1. electrons go thru CII if derived from succinate 2. heme b group (btwn C & D subunits)protects against formation of reactive oxygen species (ROS) iii. other electrons – G3P flavoprotein (G3Pdehydrogenase) on outer face of inner mitochondrial membrane Q b. Q reduced QH : m2bile electron/proton carrier i. passes electrons CIII ii. electrons from betaoxidation of fatty acids also enter here c. CIII cytochrome c (mobile connecting link) d. cytochrome c is reduced; passes electrons CIV e. CIV passes electrons O (fi2 l electron acceptor) O 2 reduced to H O 2 ALSO CIII coordinates Q cycle coenzyme QH CIII2 nding at P side of membrane intermembrane space. QH 2ives 1 electron Rieske FeS center o FeS transfers of the electron to cytochrome c , becoming reoxidized 1 o Electons passed from cyctrochome c cytoc1rome c (out of complex o QH be2omes semiquinone radical after losing that electron second electron transferred from semiquinone cytochrome b (heme b L cytochLome b H(heme b )H another CoQ bound at N side of membranematrix o CoQ fully oxidized upon passing 2 electron b cytochromes o CoQ may then dissociate from its P side binding site + 2H pumped out to intermembrane space 4. Understand how the proton gradient is generated between the matrix and the intermembrane space. Know the difference between the P side and the N side of the membrane. a. energy of electron transfer conserved in proton gradient i. transfer of 2 electrons from NADH oxygen creates LOTS of energy, used to pump protons out of the matrix ii. 1 pair electrons transferred to O 2 10 protons pumped out 1. 4 by CI, 4 by CIII, 2 by CIV iii. electrochemical energy gradient serves as temporary storage for energy of highly exergonic electron transfer b. P side (Positive side): intermembrane space + i. higher [ H ] c. N side (Negative side): matrix i. lower [ H ] 5. Understand what makes up the protonmotive force. a. 2 components net effect: ΔG 1. chemical potential energy: ΔpH i. difference in [H ] btwn 2 regions separated by membrane (IMS & matrix) 2. electrical potential energy: Δ ψ i. charge separation when proton crosses membrane w/o counteranion 6. Know the basic structure of the H ATP synthase. Understand how protons move through the protein and how ATP is synthesized. Understand the interplay between the F base and the F 0 1 head group F 1head + F 0ase + proton channel Key Asp resiue coordinates Gamma “ stick” btwn channel and head Turns w/ proton movement—transmits movement Rotates through all 3 dimers Alphabeta pair forms enzyme active site Nucleotide active site on beta ADP + P iATP Gammasubunit turning Alpha helices interact w/ alphabeta pairs Each helix has different face Face contact interaction: several configurationscyclical Open loose tight open Empty ADP +P bi nd ATP formed ATP released Study Guide for Photosynthesis – Chapter 19 1. Know the structural details of leaf and chloroplast anatomy including the unique features of each. Outer inner a. Epidermis: outermost layer b. Palisade parenchyma: “powerhouses” filled w/ chloroplasts c. Vascular system i. Xylem: water transport ii. Phloem: sugar transport d. Spongey mesophyll: only site of gas exchange & water loss i. Rest of plant has waterproof wax coating ii. CO 2iffuses in; O 2iffuses out iii. Pore = stomata e. Maximize # light photons captured 2. Understand why and how light energy is absorbed by chloroplasts and used to power photosynthesis. a. Why: light energy drives photosynthesis via the creation of electron acceptors / donors i. Complex excites electrons, which, at their higherenergy state, can be picked off by ETC b. How: 5step process i. Light photons excite light harvesting/ antenna molecule (chlorophyll or accessory pigment 1. Electron raised to higher level ii. Excitation transfer: excited antennae molecule passes energy to neighboring chlorophyll molecule, exciting it iii. Energy transferred to rxncenter clorophyll iv. Excited reactioncenter chlorophyll passes an electron to an electron acceptor v. Electron hole in rxn center filled by electron from electron donor vi. Absorption of photon separation of charge in rxn center electrochem gradient energy/ ATP to power photosythesis 3. Know the major accessory pigments, their functions, relative abundance and absorption spectra. 4 main plant photopigments (in order of relative abundance): a. chlorophyll a: green i. function: most important visible light absorption/ harvesting ii. absorption—approx peak: 665 nm and 465 nm 1. slightly better absorption than b iii. 2x more a than b in most plants b. chlorophyll b: green i. function: visible light absorption/ harvesting ii. absorption—approx peak: 640 nm (red) and 450 nm (violet blue) c. lutein (xanthophyll): yellow; type of carotenoid i. function: protects plant from excess light ( damaging radical formation) 1. radiates out extra light/ energy 2. assists chlorophylls w/ light absorption ii. absorption spectrum: 400530 nm d. betacarotene: redorange isoprenoid; type of carotenoid i. function: protects plant from excess light ( damaging radical formation) 1. radiates out extra light/ energy a. secondary protector (xanthophyll = primary) 2. assists chlorophylls w/ light absorption ii. absorption spectrum: 400500 nm (bluegreen in visible light spectrum; also some absorption in UV region) e. phycoerythrin: found more in bacteria/ algeae, lower plants i. absorbs lower light 4. Know the proteins and cofactors / prosthetic groups involved in photosynthetic electron transport. 1. Large protein complexes embedded in thylakoid membrane: ETC a. PSI i. PSI + FD + FNR + NADPH ii. Several peripheral membrane proteins attached to stromal side (see 4) b. PSII i. Associates w/ watersplitting enzyme ii. Proton gradient built btwn (outside) stroma and lumen iii. Want all energy left in stroma (for Calvin Cycle) c. Cytochrome b 6f 2. Mobile carriers a. plastoquinone (PQ)—hydrophobic (2electron carrier) b. plastocyanin (PC)—hydrophilic (1electron carrier) 3. protonATPase ATP (from gradient) 4. peripheral membrane proteins attached to stromal side of PSI a. ferrodoxin (FD) b. ferrodoxin NAD reductase (FNR) 5. FeS protens, Cu proteins/ ions a. help perform electron transport (similar to OP) 5. Understand and be able to describe electron flow through the proteins / cofactors / molecules involved in the light reactions. a. Cyclic flow i. PSI (P700) CARRIERS ferredoxin cytochrome b f complex 6 PC PSI again ii. Accompanied by proton pumping by cytochrome b f complex6 b. Noncyclic flow: “Z scheme” 1. Electrons=from H O 2SII (P680) 2. Pheo PQ AQ CytB hrome b f complex6 3. PC PSI (P700) electron acceptor chlorophyll (A ) 0 4. Phylloquinone (A ) 1FeS FeS cluster 5. ferredoxin (Fd) flavoprotein frredoxin: NADP oxidoreductase (Fd ) NADP + red + 6. (NADP reduced NADPH) 6. Know the major products of the light reactions and the Calvin cycle and the locations of these two processes. a. Light reactions: i. Products: ATP, NADPH, O 2 ii. Location: thylakoid membrane of chloroplasts b. Calvin cycle (a.k.a. Lightindependent reactions): I + i. Products: ADP + P, NADP , sugars (sucrose or starch) 1. Sucrose: synthesized in cytoplasm 2. Startch: stored/ produced in chloroplast (no transport) ii. Location: stroma of chloroplast 7. Understand how the watersplitting complex of PS II works. a. Watersplitting enzyme has 2 parts i. Mn cluster w/ calcium ion= active site—splits water ii. Tyr residue—KEY iii. Intermediate btwn enzyme complex & P680 b. P680 resets self by immediately “stealing” electron from Tyr i. Tyr radical; not favorable ii. Tyr stabilizes by stealing electron from Mn iii. Mn stable from 2+ 4+ so OK c. Electronstealing process happens 4 times th i. Each time, Mn becomes stronger oxidizer (strongest after 4 excitation) ii. Capable of splitting 2 H20 molecules iii. PSII Takes back 4 electrons to reset itself 8. Know the distribution of the photosynthetic machinery on the thylakoid membranes. Left right PSII (P680) Q cycle (PQ/ PQH ) Cytochrome b f PC PSI (P700) ATP 2 6 synthetase PSII: thylakoid stacks PSI in lamellae Cytochrome b f m6stly in lumen 9. Understand how the proton gradient is generated between the lumen and the stroma. Know the difference between the P side and the N side of the membrane. a. How: VERY similar mechanism to mitochondria i. CF like mitochondrial F : transmembrane proton pore composed of 0 0 several integral membrane proteins ii. CF l1 e mitochondrial F : pe1ipheral membrane protein complex iii. ATP synthase complexes on the outside of thylakoid membranes (N stromal side)—opposite of complexes on inside of mitochondria (N matrix side) 1. iv. ADP & P condense on enzyme surface release of enzymebound ATP i requires protonmotive force v. Rotational catalysis engages all 3 beta subunits of ATP synthase 1. ATP synthesis subunit 2. ATP release subunit 3. ADP + P biniing subunit b. P side (Positive side): lumen i. higher [ H ] c. N side (Negative side): stroma i. lower [ H ]+ 10. Understand basic information about the Calvin cycle (READ Chapter 20 pages 799809) such as: a. Location of Calvin Cycle i. Stroma of chloroplast b. Enzyme that catalyzes initial carboxylation/fixation reaction c. Three stages of the Calvin cycle 1. CO fixation 2 2. Reduction 3PGA G3P 3. G3P RuBP (regeneration) d. Initial acceptor molecule that must be regenerated i. RuBP e. Sugar precursor that is formed i. G3P Study Guide for Comprehensive portion of the Final Exam – BSC 450/550 Chapter 2: 1. Use the HendersonHaselbach equation to solve problems. 2. Be able to interpret titration curves of weak acids and bases i.e. areas of greatest and least buffering, etc. Reveals pk_a (at center) 3. Understand how buffers work and what buffering capacity means. a. Buffers added to solution to reduce the increase of acicidity or basicity as acid or base is added (i.e. change in pH) b. Buffering capacity: pH range at which buffer is able to resist/ minimize pH change i. Flatter portion near middle of titration curve ii. Works best w/n 2 pH units of pH of original solution (+/ 1 unit) iii. Ex. for blood pH = 7.4, the range would be 6.48.4 iv. Max buffering capacity: pH = pk_a Chapter 3: 1. Know the structure, three letter code and single letter code for the amino acids. • Aliphatic: Ala (A), Val (V), Leu (L), Ile (I), Met (M), Pro (P), Gly (G) • Nonpolar, aromatic: Phe (F), Tyr (Y), Trp (W) • Polar, uncharged: Cys (C), Ser (S), Thr (T), Asn (N), Gln (Q) • Polar, positively charged: His (H), Arg (R), Lys (K) • Polar, negatively charged: Asp (D), Glu (E) OR Hydrophobic (aliphatic): Ala (A), Val (A), Leu (L), Ile (I) Hydroxylcontaining: Ser (S), Thr (T) Aromatic: Tryptophan (W), Tyrosine (Y), Phenylalanine (F) SulfurContaining: Cys (C), Met (M) Outlier: Pro (P), His (H) Acidic: Asp (D), Glu (E) Amidecontaining: Asn (N), Gln (Q) Basic: Lys (K), Arg (R) 2. Be able to interpret titration curves of amino acids and calculate pI. • pI = average of pk’s on either side of zwitterionic state of aa • zwitterion: net charge = 0 • sum of charges on carboxyl group, amino group, and side chain Chapter 6 1. Be able to determine K andmV frommaxta in a table or on a plot. a. V max : highest value of v b. K :m[S] corresponding to half that value c. K =m K + K1 / K2 = [S1 @ V = .5 x0V max i. K 1 describes ES formation ii. K 1describes ES breakdown iii. k2 : describes product formation (irreversible, ratedetermining) d. v = maximum velocity (plateau of hyperbola on Michaelis plot) as [S] is m increased i. reached when all enzyme activates sites are filled with substrate, forming ES complex (doesn’t count if it’s an ESI complex) e. k m V max slope of LineweaverBurk plot f. decreasing k mbetter enzyme i. less substrate needed to get to same velocity g. increasing v better enzyme max h. k /cat =mspecificity constant: good measure of enzyme efficiency i. “catalytic perfection”: specificity constant 10 10 M S9 1 1 2. Understand and be able to recognize the different enzyme kinetic mechanisms (sequential vs. pingpong). a. Sequential: substrates bond to enzyme concurrently, forming noncovalent ternary complex i. Random: substrates bind in either order ii. Ordered: necessary that 1 binds for the next to bind efficiently b. Pingpong (i.e. double displacement): first substrate converted to product before 2 substrate binds to transformed enzyme; release of 2 enzyme regenerates original enzyme; no ternarny complex i. Denoted by 2 parallel slopes on LineweaverBurke plot 3. Be able to identify the different types of enzyme inhibition. a. Irreversible: permanent shutdown i. Suicide (i.e. mechanismbased): irreversible covalent bonding; hijack enzyme’s normal mechanism to produce inactive form; useful in rational drug design ii. Transitionstate analogs: very tight noncovalent binding; Resembles transition states; fits enzyme active site better than substrate b. Reversible i. Competitive: bind to active site of E ii.Uncompetitive: bind to TS on another site once ES complex has formed iii. Noncompetitive (mixed): binds at site that isn’t the active site; can bind to either E or ES 4. Be able to recognize different mechanisms of enzyme kinetics and enzyme inhibition from LineweaverBurk plots. a. Enzyme inhibition i. Competitive: inhibits substrate binding; no effect on catalysis 1. No change v ; maxrease in k m 2. no change in k cat 3. slopes intersect at yaxis ii. Uncompetitive: inhibits catalysis; does not affect substrate binding 1. Decrease in v max decrease in k m; change v max / m 2. No change in k cat 3. parallel slopes iii. Noncompetitive/mixed: inhibits substrate binding and catalysis 1. Decrease in v max no change in k m; 2. Reduced k cat 3. slopes intersect left from yaxis b. Enzyme kinetics i. intersection of slopes at single point: ternary complex 1. ordered or random 2. intersection away from yaxis ii. parallel slopes: pingpong (double displacement) Chapter 7 1. Understand carbohydrate naming (triose, ketone, aldehyde, etc.) and vocabulary (epimers, anomers, pyranose). a. Aldo/keto + __prefix__ + “ose” i. Prefix: triose: 3C; tetrose: 4c; pentose: 5C; hexose: 6c, heptose: 7C… ii. Epimer: 1 of 2 isomers with different configurations around 1 asymmetrical C iii. Anomer: specific type of epimer; 1 of 2 stereoisomers w/ differing configuration only at hemiacetal/acetal carbon (a.k.a. anomeric carbon) iv. Pyranose: 6membered carbohydrate ring w/ 5 C’s, 1 O v. Furanose: 5membered carbohydrate ring 2/ 4 C’s, 1 O 2. Be able to name glycosidic linkages. a. Determine configuration of each sugar, prelinkage i. When linear: L v. D; look @ C furthest from carbonyl C 1. –OH on left = L; OH on right = D ii. Alpha v. beta: look at anomeric C 1. Alpha: anomeric –OH on opposite side of –OH on C6 2. Beta: anomeric –OH on same side of –OH on C6 b. Name left right; nonreducing reducing i. 1. Nonreducing configuration (L/Dalpha/beta) + 2. Nonreducing name, cutting off at “o” + furanosyl / puranosyl + 3. “(#C of original hemiacetal #C of original alcohol)” (most common 14) + 4. Reducing configuration + 5. Reducing name Chapter 8 1. Understand the various molecular mechanisms responsible for DNA mutagenesis a. Oxidative (more easily repaired by cellular processes) i. Spontaneous 1. Deamination a. Slow, daily process (100 CU per day) 2. Depurination a. Hydrolysis of betaNglycosyl bond (loss base, form AP site) b. Significant for purines (10k lost per day) c. Repaired by baseexcision repair ii. Reactive chemicals 1. Oxidation by hydroxyl radical a. Hydroxylation of guanine b. Mitochondrial DNA most susceptible 2. Methylation of guanine (on carbonyl O) b. Radiation i. UV (dimerization of thymidine) 1. Main mechanism for skin cancer ii. Xray (break covalent bonds ring opening, strand breaking ) 1. Ionizing 2. Worst kind; most difficult to repair 2. Understand why the inclusion of thymine in DNA only was an important evolutionary leap a. “better protection” of DNA storage molecule i. spontaneous deamination transforms C U 1. genetic mutation high A=T : G=C ratio 2. example of spontaneous oxidation damage to DNA a. easiest to repair ii. repair enzyme: uracil DNA glycosylase 1. can safely excise U bases from doublestranded DNA formed due to this process, bc they should never be there 2. indicates problem; initiates repair 3. would not be able to do this in RNA, where U bases are legitimately present b. more important to have this feature in DNA, bc mRNA is shortlived but DNA errors are passed on in every replication Chapter 10: 1. Know the basic functions and structures of the lipid soluble vitamins A, D, E and K. a. Vitamin A: vision (particularly light sensitivity and night vision) i. Rods and cones: cis trans configuration when dark light b. Vitamin D: calcium uptake i. Activated in 2 steps in skin by UV light ii. Cholecalciferol (Vit D3): activated inside the body c. Vitamin E: antioxidant i. Interact w/ free radicals to inactivate them quenching ii. more prevalent in mitochondria due to greater oxidation susceptibility d. Vitamin K: essential to blood clotting; activates prothrombin Chapter 11: 1. Understand the concept of lipid rafts, how they are formed and why they are important. a. Formed in membrane from cholesterolsphingolipid association b. A. k. a. microdomains: thicker and more ordered than surrounding monolayer of PM leaflets c. Enriched by 2 classes of integral membrane proteins i. GPIanchored proteins (outer) ii. Cysattached proteins w/ 2 longchain saturated FA’s (inner) d. vesicle formation – important for endocytosis i. caveolin (ex. Cysattached) recruits claithrin endocytosis e. role in signaling 2. Understand the mechanisms behind the ability to curve the cell membrane, form vesicles and fuse them to other membranes. a. Fusion i. **Sperm + egg ii. Endosomes + lysosomes iii. Small vacuoles larger vacuoles (plants only) b. Membrane curvature i. Vesicle formation (budding) from Golgi complex ii. Exocytosis iii. Endocytosis 1. Mechanism of viral infection iv. **cell division—separation of 2 plasma membranes c. Both: neurotransmitter release Chapter 12: 1. Describe the six basic types of signal transducers in the cell. a. G proteincoupled receptor (GPCR) –see #6a for more detail i. External ligand binds to receptor intracellular GTPbinding protein activation ii. Activation regulation of enzyme that generates intracellular 2 d messenger iii. Beststudied receptor: targets 50% of drugs 1. Ex. beta blocker: suppresses fight/flight response reduced anxiety b. Receptor tyrosine kinase: ligand binding tyrosine kinase activity (autophosphorylation) i. Kinase transcription factor alteration altered gene expression ii. Predominant in animals; Ser/Thr more common in plants c. Receptor guanylyl cyclase: ligand binding to extracellular domain formation nd cGMP (2 messenger) d. Gated ion channel: opens/closes based on signal ligand concentration or membrane potential i. Specific response to extracellular ligand ii. Simplest type of bonding e. Adhesion receptor (integrin): binding molecules in ECM altered conformation changed interaction w/ cytoskeleton i. tails for clamping onto cytoskeleton ii. physical transduction across membrane 1. bent = off; straight = on –recruits adapters f. Nuclear receptor: hormone binding regulation of specific genes 2. Describe the five factors that contribute to the sensitivity / unique properties of signal transduction pathways a. Specificity: ONLY particular signaling molecule fits complementary binding site on receptor b. Amplification: enzymes affecting enzymes geometric increase in affected molecules enzyme cascade c. Modularity: proteins w/ multivalent affinities form diverse signaling complexes/modules from interchangeable parts i. “multivalent affinites” = affinity for multiple signaling molecules ii. KEY for building: phosphorylation 1. Forms new binding sites 2. Provides reversible points of interaction d. Desensitization: feedback inhibition i. Activated receptor triggers shutdown of receptor or removal from cell surface ii. Expression turned “off” or “down” e. Integration: crosstalk between pathways i. Antagonistic: addition of 2 pathways w/ opposing effects on a metabolic characteristic ii. Regulatory outcome = integrated input of both pathways 1. Some cancellation 2. Net result reveals which pathway predominates iii. Responsible for generalized immune response (evolutionarily conserved) 3. Understand and be able to describe the basic structure of a Gprotein coupled receptor and its associated heterotrimeric Gprotein a. GPCR: i. 3 outer loops: bind epinephrine ii. 3 inner loops: associate with Gprotein iii. alpha unit: GDPbound: off; GTPbound: on iv. Dissociation active effector (typically, an enzyme) v. Beta unit: no lipid tail b. Gprotein: guanosinenucleotide binding protein i. Heterotrimeric: 3 subunits G alpha, Gbeta, and Ggamma s 1. G aspha: contains pocket for GDP binding a. Activated by epinephrine binding 4. Know the specific changes that occur to the β AR w2en epinephrine binds and the changes that this causes in the Gα subunit. a. Epinephrine binds to specific receptor formation hormone receptor complex b. Complex causes GTP to replace GDP activation G salpha c. Activated G salpha es from G sbetagamma adenylyl cyclase activation adenylyl cyclase i. One complex may activate many G salpha a. Binding changes in TM5 / TM6 ii. TM5 shifts down iii. New loop formed in TM6 (new 2ndary structure) b. Alpha subunit on G protein intimately associated w/ receptor; change shape receptor new interaction w/ alpha subunit c. TM5 shift new hydrophobic pocket created so G alphaan shift up d. F139: phenylalanine iv. critical in messagement movement v. without it, epinephrine still binds but no signaling https://studysoup.com/universityofalabamatuscaloosa/bsc450/study guide/bsc450finalexamstudyguide?id=94961
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