Study Guide

Ch. 1.  noncovalent interactions: hydrogen bonds, electrostatic bonds, van der Waals interactions; be able to do calculations with pH, Henderson-Hasselbalch equation; buffers.

Ch. 2.  Be able to draw the structures of the 20 amino acids at pH 7.0; know the three-letter codes and one-letter codes for the amino acids.  Know which ones are acidic, basic, hydrophilic, and hydrophobic. L stereoisomers.  Be able to use the Henderson-Hasselbalch equation in calculations on amino acids. Know the levels of proteins structure.  Structure of the peptide bond, peptide nomenclature, N- and C-terminal; disulfide bond.  Native protein; protein folding is driven by burying hydrophobic residues in interior and by maximizing the number of hydrogen bonds; peptide bond; primary structure = amino acid sequence; secondary structure:  a helix (what amino acids stabilize or destabilize), b pleated sheet (parallel and antiparallel); tertiary structure:  prosthetic group, supersecondary structure, motifs (helix-turn-helix), domains; protein denaturation by guanidinium, urea, b-mercaptoethanol; primary structure determines tertiary structure:  Anfinsen experiment.

Ch. 3  purification of proteins:  choose an abundant source, assay.  Column chromatography:  ion-exchange, size-exclusion, affinity chromatography; calculation of yield and specific activity to monitor protein purification; SDS PAGE, log molecular weight vs. relative mobility plot can be used to determine the molecular wt. of the subunits of a protein, SDS gels can monitor purification of a protein.  Sequencing:  Edman degradation (with PITC); longer peptides are specifically cleaved with trypsin, chymotrypsin, or CNBr (know the specificities for these); protein sequence can be deduced  from DNA sequence.  Antibodies, immunoglobulins, IgG; structure of IgG (Fig. 3.27), immunoglobulin fold (3.27); antigenic determinant, epitope, complementarity-determining region (CDR); polyclonal antibodies, monoclonal antibodies (Fig. 3.30), herceptin; Fig. 3.32 indirect ELISA and sandwich ELISA, ELISA kit for hCG, Western blot (immunoblot) (Fig. 3.33).

Ch. 11.  Monosaccharides:  all sugars are D-sugars, aldose, ketose, recognize structure of glucose, fructose, and ribose; anomeric carbon, be able to recognize the structure of a and b anomers, chair form of glucose is most stable; N- and O-glycosidic bond; Fig. 11.9 names and abbreviations of modified monosaccharides; composition of sucrose, lactose, and maltose; structure of glycogen, amylose, amylopectin, and cellulose; recognize the names of glycosaminoglycans and that they are components of proteoglycans (example:  aggrecan), part of connective tissue and extracellular matrix; glycosyltransferases and their importance in human A, B, and O blood types; N-linked (asn) and O-linked (ser, thr) carbohydrate; Lectins:  selectins bind immune cells to sites of injury by bindng carbohydrate Leukocyte rolling and extravasation; influenza hemagglutinin binds to sialic acid residues on cell-surface glycoproteins.

Inner life of a cell, including leukocyte extravasation:

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Ch. 4, 28 Be able to draw the structures of all the nucleosides and nucleotides and be familiar with the nomenclature; phosphodiester linkage; Watson-Crick structure of DNA Fig. 4.11; base pairs Fig. 4.12, major groove, minor groove Figs. 28.5 and 28.6, B-DNA and A-DNA Fig. 28.3; DNA denaturation and annealing, Fig. 4.17.  DNA replication is semiconservative, bidirectional (at 2 replication forks) from an origin of replication, 5’-3’ direction, leading strand, lagging strand and Okazaki fragments; Fig. 4.21, DNA polymerases require a template, primer with free 3’ OH, 4 dNTPs; all DNA polymerases have 3’-5’ exonuclease proofreading activity; processivity, DNA polymerase I has 5’-3’ nuclease activity, nick translation; the b subunit of DNA polymerase III is responsible for its high processivity; replication fork:  helicase, ssDNA-binding proteins, primase, DNA polymerase III (a, b, e subunits and their functions), clamp loading complex , DNA polymerase I, DNA ligase (know what each of these does in DNA replication); initiation is at oriC; elongation Fig. 28.29; termination at Ter sequences with Tus protein; characteristics of eukaryotic DNA replication; telomerase replicates eukaryotic chromosome ends Fig. 28.34; cytosine deamination repair Fig. 28.45; reverse transcriptase of retroviruses Fig. 4.23.

Ch. 29 RNA structure; mRNA, rRNA, tRNA, snRNA, miRNA and si RNA; RNA polymerases need a template and 4 NTPs but no primer, synthesis is in 5’-3’direction; template and nontemplate (coding) strands; function of s subunit of RNA polymerase; transcription start site = promoter, the RNA polymerase binding site; components of prokaryotic promoter; a promoter sequence identical to the consensus sequence is a strong promoter; termination signals for prokaryotic transcription; eukaryotic RNA polymerase II, promoter:  TATA box, Inr, DPE, CAAT, GC, Enhancer; TBP, TFIIs are required for the binding of RNA polymerase II to the promoter; mRNA processing in eukaryotes:  5’ cap, poly A tail, splicing; know function of snRNA and snRNPs in splicing; ribozyme; complex transcripts:  alternative splicing; RNA editing.

Ch. 30 structure of tRNAs; aminoacyl tRNA synthetases (use ATP hydrolysis to charge tRNAs with amino acid);;  the ribosome (50S + 30S = 70S) is composed of rRNA and protein:  the 16S rRNA aligns the ribosome with the Shine-Dalgarno sequence in the mRNA for initiation of translation; the 23S rRNA catalyzes the formation of the peptide bond (a ribozyme); initiating tRNAs for bacteria and eukaryotes; direction of synthesis is N to C, 5' to 3' along mRNA; Shine-Dalgarno sequence; initiation:  initiation factors, GTP hydrolyzed (Fig. 30.22); elongation:  EF-Tu, EF-Ts, GTP hydrolyzed (30.19); peptide bond formation (30.19); translocation:  EF-G, GTP hydrolyzed (30.24); codon, reading frame, initiation codon (AUG), termination codons (UAA, UAG, UGA), Wobble hypothesis; termination:  release factors (30.25); eukaryotic initiation (30.26)

EXAM 2 STARTS HERE:

Ch. 5 restriction endonucleases: palindromic recognition sites, blunt or staggered cleavage (sticky ends); Sanger method of DNA sequencing (p. 138); PCR (5.8); DNA fingerprinting  using STRs; Recombinant DNA (5.11) DNA ligase; polylinker; vectors: plasmids (5.12 and 5.13), bacteriophage l (5.15) transform host by infection, l replication; YACs (5.16), yeast chromosome replication  for each of the vectors have a ballpark estimate of the size of foreign DNA they can accommodate, how to transform cells, and how they replicate in the host cell; genomic library (5.17); expression vectors; cDNA library (made from tissue mRNAs using reverse transcriptase) (5.28 and 5.29); RNAi and siRNA, (5.36).

Ch. 7  recognize structure of heme (Fe²⁺ protoporphyrin IX) ; O₂ binding to myoglobin (fig. 7.4), hyperbolic O₂ binding curve (Fig. 7.6) ;  hemoglobin structure: a₂b₂, sigmoidal O₂ binding curve (Fig. 7.7), isolated b chain has structure and O₂ binding like myoglobin, hemoglobin carries O₂, CO₂, and H⁺; cooperativity, T state and R state, T state is stabilized by electrostatic interactions between subunits (in deoxyHb, Fe is pulled out of the plane of the heme, Fig. 7.14) BPG, O₂ binding to fetal hemoglobin; electrostatic bonds stabilize T state:  BPG, CO₂, and H⁺ (Bohr effect) as heterotropic allosteric effectors of hemoglobin

Ch. 8.  cofactor, coenzyme, prosthetic group, holoenzyme, apoenzyme; memorize and be able to do calculations with Eq. 1 on p. 209; enzymes are catalysts which alter the rates of the reaction they catalyze but not the equilibrium between substrates and products or the free energy available from the reaction; transition state, activation energy; induced fit; how do enzymes catalyze reactions?  the enzyme binds the substrate in the shape of the transition state to lower the activation energy; active site; memorize the Michaelis-Menten equation, V_max, K_m; be able to derive V_max and K_m from a V vs. [S] plot or a Lineweaver-Burk plot, be able to calculate turnover number.  

Reversible inhibition:  competitive inhibition:  inhibitor often resembles the substrate, it binds at the active site, competitive inhibition can be overcome at high substrate concentration; know what V vs. [S] plot and Lineweaver-Burk plot for competitive inhibition look like.  Irreversible inhibitors: acetylcholinesterase, suicide inhibitors; transition-state analogs as enzyme inhibitors (amoxicillin/clavulanate).

 

Ch. 10. allosteric:  first enzyme in a pathway, usually more than one subunit, sigmoidal V vs. [S] plot, allosteric activators and inhibitors, aspartate transcarbamoylase as model (ATP, CTP); isozymes:  lactate dehydrogenase as example); reversible covalent modification:  example is phosphorylation by serine/threonine or tyrosine kinases and reversible by phosphatases; proteolytic activation, zymogens and proproteins (chymotrypsin example).

Ch. 12. Be able to draw a fatty acid given the number of carbons and double bonds and be able to give the carbon number and double bond location for a fatty acid (Table 12.1); understand the 2 ways of numbering fatty acids and what w-3 and w-6 signify;   fatty acids contain cis double bonds;  length and unsaturation determine melting temperature of a fatty acid; recognize structure of membrane lipids and be able to describe their components (eg., Fig. 12.4, 12.6, and p. 331 cerebroside) ; recognize glycerophospholipids in Fig. 12.5 ; recognize ceramide, sphingomyelin (12.6), glycolipids (cerebrosides p. 331 and gangliosides); cholesterol (p. 331).  Structure of a lipid bilayer (Fig. 12.11) and liposomes (Fig. 12.12); nonpolar molecules can move through the bilayer rapidly, polar and charged species move slowly and require protein transporters; lateral and transverse mobility of membrane lipids (Fig. 12.31) and proteins; structure of membrane proteins (Fig. 12.27), carbohydrate is attached on the outside surface of the cell; peripheral and integral membrane proteins (Fig. 12.17); hydropathy plots (Fig. 12.27 and Table 12.2); most membrane proteins are made of a helices (Fig. 12.18); b barrel motifs are found in porins (Fig. 12.20); lipid anchors for membrane proteins, with farnesyl and GPI examples (Fig. 12.26).  

Ch. 13. uniport, symport, antiport; active vs. passive transport; memorize the equation and be able to calculate DG for transport (p. 353); carriers and channels; kinetics of carrier-type transport and understand the significance of the different K_ts for glucose transport for GLUT1 and GLUT2; active transport:  pumps:  Na⁺, K⁺ ATPase, Ca²⁺ ATPase (SERCA) mechanism (Fig. 13.5); Na⁺- and H⁺-driven secondary active transport (example, lactose-proton symport by lactose permease Fig. 13.12); channels catalyze passive transport of ions;  potassium channel structure (Fig. 13.17 and 13.19 selectivity filter); voltage-gated K⁺ channels (Fig. 13.22 and 13.23):  mechanism of voltage gate, ion selectivity mechanism, and inactivation (Fig. 13.25); ligand-gated (structure and mechanism of acetylcholine receptor (Fig. 13.26, 13.28, and class notes)); mechanism of the action potential Fig. 13.30.

 

EXAM III STARTS HERE:

Ch. 14.  G protein-coupled receptors (GPCR or 7TM) with b-adrenergic receptor as example (Fig. 14.5) cAMP activates protein kinase A; ; off switches:  Figs. 14.9, 14.10.  insulin receptor tyrosine kinase Fig. 14.20, pleckstrin homology domain, SH2 domain, movement of GLUT 4 to the plasma membrane, and termination by phosphatases. YOU SHOULD KNOW NO SYNTHASE MAKES NO FROM ARGININE.  NO ACTIVATES GUANYLATE CYCLASE, WHICH MAKES cGMP.  cGMP IS INACTIVATED BY cGMP PHOSPHODIESTERASE. Defects in signal transduction and cancer:  v-Src, Ras, EGFR (Herceptin).

 

Ch. 15.  ATP hydrolysis drives many cell reactions (eqns. at the top of p. 413); catabolism (Fig. 15.12), anabolism; recognize NAD⁺/NADH, NADP⁺/NADPH, FAD/FADH₂, coenzyme A

 

Ch. 16.  Fig. 16.2:  know names of glycolytic intermediates and be able to write out the complete pathway, where ATP is consumed and formed, where NADH is formed, and net reaction; controlling enzymes: hexokinase, phosphofructokinase-1, pyruvate kinase.  Pyruvate can be converted to ethanol, lactate or CO₂ to regenerate NAD⁺.  Location of glycolysis in cell.  Fig. 16.21:  tumors use glycolysis for energy.

Gluconeogenesis makes glucose from the noncarbohydrate precursors lactate, glycerol, propionate, and amino acids.  Located in the mitochondrion, cytosol, and endoplasmic reticulum; know which enzymes in gluconeogenesis bypass the irreversible steps of glycolysis (Fig. 16.22); pyruvate carboxylase contains biotin and is a regulatory enzyme; stoichiometry of gluconeogenesis (Eqn. on p. 464).

 

Ch. 17.  pyruvate dehydrogenase complex:  eqn on p. 477, be able to recognize thiamine pyrophosphate, lipoamide, FAD, it is a regulatory enzyme. 

citric acid cycle:  acetyl CoA is oxidized to 2 CO₂ with the formation of 3 NADH, 1 FADH₂, 1 GTP;  intermediates in the citric acid cycle are used to synthesize other biological molecules; be able to name all the intermediates in the citric acid cycle in order and know at which steps NADH, FADH₂, GTP, and CO₂ are formed (Fig. 17.15); regulatory enzymes are citrate synthase, isocitrate dehydrogenase, and a-ketoglutarate dehydrogenase.

anaplerotic reactions (example, pyruvate carboxylase reaction).  Recognize structure and know function of biotin.

 

Ch. 18  mitochondrion structure; be able to identify the structures of ubiquinone, FMN, hemes of the cytochromes, and iron sulfur centers; Fig. 18.17:  reactions of mitochondrial electron transport, names of complexes and their order, Q, and cytochrome c, proton pumping by complexes I, III, and IV; given DG = RTln c₂/c₁ + zFDy be able to calculate DG; chemiosmosis:  proton pumping drives ATP synthesis; structure of ATP synthase and mechanism (Fig. 18.29 and 18.32); thermogenin; superoxide and peroxide are reactive species produced by the metabolism of O₂ and are removed by superoxide dismutase and catalase, respectively.

 

Ch. 19 structure of chloroplast; recognize chlorophyll;lin photosynthetic electron transport, water is oxidized to O₂ and NADPH and ATP are formed; components of the photosynthetic electron transport in order including H₂O, oxygen-evolving complex, photosystem II, plastoquinone, cytochrome bf complex, plastocyanin, photosystem I, ferredoxin, ferredoxin-NADP⁺ reductase, NADP⁺;  Fig. 19.25.  

 

Ch. 20  Fig. 20.1 the Calvin cycle, located in chloroplast stroma; rubisco is the regulatory enzyme and has carboxylase and oxygenase activities; phosphoglycolate from oxygenase activity must be salvaged; C₄ plants incorporate CO₂ into C₄ intermediates in the mesophyll cells, these are transported to bundle sheath cells where they generate CO₂ for the Calvin cycle there, and this minimizes the oxygenase activity of rubisco (Fig. 20.17).

Pentose phosphate pathway: located in cytosol, glucose 6-phosphate dehydrogenase is regulatory enzyme;  oxidative branch:  glucose 6-phosphate is converted to ribose 5-phosphate and 2 NADPH are produced; nonoxidative branch: 3 C₅ intermediates are converted to 2 C₆ and C₃.  

 

Ch. 21 glycogen degradation:  located in cytosol; be able to describe the structure of glycogen; degradation:  glycogen phosphorylase (regulatory enzyme) (phosphorolytic cleavage) and debranching enzyme and phosphoglucomutase.  glycogen synthesis:  located in cytosol; UDP-glucose is "activated" form of glucose used for synthesis; glycogen synthase (regulatory enzyme); branching enzyme; glycogenin.  Regulation of glycogen phosphorylase:  GPCR regulation:  Fig. 21.17.

 

Ch. 22 Fig. 22.6:  mobilization of stored triacylglycerols:  glucagon and epinephrine activate triacylglycerol lipase using cAMP as a second messenger, perilipin; glycerol can be converted to DHAP for gluconeogenesis; serum albumin

fatty acids are attached to CoA in the cell; carnitine acyltransferase I is the regulatory enzyme for fatty acid oxidation; fatty acids are transported into the mitochondrion as carnitine derivatives.

b oxidation:  during each round of oxidation, 1 acetyl CoA, 1 FADH₂, and 1 NADH are formed; acetyl CoA is oxidized to 2 CO₂ in the citric acid cycle; odd-chain fatty acids are oxidized until propionyl CoA is left-the propionyl CoA is converted to succinyl CoA in a biotin- and coenzyme B₁₂-dependent pathway; recognize coenzyme B₁₂

ketone bodies:  acetoacetate, b-hydroxybutyrate, and acetone are produced from acetyl CoA;  they can be used as fuel by skeletal muscle, heart, renal cortex, and (during starvation) the brain; Fig. 22.21 and 22.23

Fatty acid synthesis:  located in the cytosol, acetyl CoA carboxylase is the regulatory enzyme and contains biotin; malonyl CoA is used in synthesis, fatty acid synthase, intermediates are carried on acyl carrier protein (ACP), 2 NADPH required per round; to make palmitate, need 8 acetyl CoA, 7 ATP, and 14 NADPH; citrate carries acetate groups (8) to cytosol, and this process generates 8 NADPH.  

.   Exam III ends here.

The following lectures will be on the final exam.

Ch. 23  Fig. 23.1.  Ubiquitin tags proteins for destruction, lysines in target proteins form an isopeptide bond with the C-terminus of ubiquitin; E1, E2, and E3 catalyze this reaction; E3 detects the N-terminal amino acid of the target protein, which determines the protein’s half life; cyclin destruction boxes and PEST sequences are other destruction signals.  Structure of the proteasome Fig. 23.6, polyubiquitinated proteins are degraded by the proteasome; bortezomib inhibits the proteasome and is used to treat multiple myeloma since it inhibits destruction of pro-apoptotic proteins.  Know the alanine aminotransferase and aspartate aminotransferase reactions; pyridoxal phosphate is the prosthetic group (it is involved in many reactions of amino acid chemistry:  aminotransferases, racemases, decarboxylases, aldolases), be able to recognize its structure (Fig. on p. 657). Fig. 23.16, glutamine synthetase and glutaminase reactions.  Urea cycle:  Fig. 23.17 and where ATP is used.  Fig. 23.22:  know the definition of glucogenic and ketogenic amino acids, know one amino acid for each intermediate.  Phenylalanine hydroxylase deficiency results in phenylketonuria, it is detected in newborns and alleviated by using a low phenylalanine diet.

Ch. 24  Rhizobium symbiosis (what the plant supplies, what the bacteroid supplies, leghemoglobin); stoichiometry of the nitrogenase complex (p. 6815); nitrogenase complex is composed of dinitrogenase reductase (Fe protein) and dinitrogenase (MoFe protein) Fig. 24.1; reactions catalyzed by gln synthetase and glu synthase; Fig. 24.5 know one amino acid for each precursor and that pentose phosphate pathway, glycolysis and citric acid cycle intermediates are used to synthesize amino acids; Fig. 24.9 tetrahydrofolate, which carries 1-C units in several oxidation states; S-adenosylmethionine p. 691 carries methyl groups; NO synthase makes NO from arginine; know the amino acids which are used to synthesize neurotransmitters and which neurotransmitters they make, and that pyridoxal phosphate is involved.

 

Ch. 25. Purine synthesis (Fig. 25.5-know where the atoms in the purine ring come from):  the base is built on the sugar (phosphoribosyl pyrophosphate, PRPP) in the cytosol; gln-PRPP amidotransferase is a regulatory enzyme.  Pyrimidine synthesis:  the base is formed and then added to PRPP in the cytosol; aspartate transcarbamoylase is the regulatory enzyme; know the source of the atoms in the pyrimidine ring; ribonucleotide reductase; thymidylate synthase pathway (Fig. 25.13).

Ch. 26. All 27 C atoms of cholesterol come from acetyl CoA; location:  cytosol,  regulatory enzyme, HMG CoA reductase; know the pathway of cholesterol synthesis by the number of carbons as shown in class, recognize the structure of isopentenyl pyrophosphate (be able to spell it right), importance of farnesylation; steroids, bile salts, and vitamin D are made from cholesterol.

Ch. 27.  For insulin and glucagon, know their effect on blood glucose, their mechanism of signal transduction; insulin stimulates storage and synthesis pathways; glucagon stimulates energy mobilization pathways; mechanism of insulin release by the pancreas in response to high blood glucose (handout and class notes). Metabolism in Type I and Type II diabetes mellitus.  Body mass regulation:   leptin is produced by adipocytes and stimulates the hypothalamus to produce anorexigenic peptides (suppress eating) and stimulates sympathetic neuron signals (see handout and class notes for norepinephrine pathway) to cause TAG breakdown and the synthesis of UCP to promote thermogenesis and fatty acid oxidation.