Chemistry 350
Spring 2000

Dept. of Chemistry and Biochemistry
Southern Illinois University at Carbondale

Objectives for Unit 1

Protein structure, Carbohydrates, Membranes

1. Write structure of any of the amino acids or peptides when given the three letter designation of the amino acid or sequence. Identify functional groups, side chains, ionic forms of amino acids; classify amino acids according to type (e.g., basic, acidic, aliphatic, etc.) and properties (e.g., charged, H-bond capacity, hydrophobicity, etc.)
2. Work all problems of the type assigned, as well as examples covered in class.
3. Be able to accurately sketch a titration curve for an amino acid or small peptide when given the pKa values. Draw the structure of the predominant species that would exist at a specified pH. Determine the approximate charge on the molecule at any point along the titration curve. Identify the ionic specie(s) that would exist at a specified pH and accurately calculate the pl.
4. When given the pl(s) or other relevant data that would let you determine the net charge on a molecule at a particular pH, predict the direction and relative rates of migration in an electrical field.
5. Determine the structure of a peptide when given sequence data. Know specificities of reagents and enzymes used in protein sequencing and be able to apply them to sequencing problems.
6. Outline an experimental procedure for determination of a peptide sequence when given a particular peptide.
7. Know reactions, tests, and reagents used in amino acid identification and quantitation.
8. Understand the significance of primary structure, secondary structure, tertiary structure, and quaternary structure.
9. Recognize a-helix and parallel and antiparallel b-sheet structures, hydrogen bonds, hydrophobic interactions, ionic (electrostatic) bonds and disulfide bonds in a diagram of a protein. Be able to describe simple motifs ( helix-loop-helix, hairpin beta, greek key) and domains.
10. Understand the importance of the marginal stability of protein molecules, describe methods for studying the progressive stabilization of intermediates in protein folding.
11. Define all terms we will have had related with amino acids, peptides, carbohydrates, and lipids.
12. Understand the principles behind the following methods used for isolation of low and high M. W. biological compounds: gel electrophoresis, SDS gel electrophoresis, dialysis, salting out, ion exchange chromatography, affinity chromatography.
13. Write open chain and cyclic structures for: D-glucose, D-fructose, D-galactose, D-ribose.
14. Draw disaccharide structures showing alpha, beta, (1-4), (1-6), etc. linkages of the above sugars.
15. Define all terms we will have had related to carbohydrates and simple sugars, know the characteristics of the most important di- and polysaccharides.
16. Be able to answer questions involving mutarotation and oxidation-reduction of sugars.
17. Know the types of lipids found in membranes and their characteristics.
18. Know the components and characteristics of the fluid mosaic model for membranes.
19. Understand the following concepts regarding membranes: peripheral and integral proteins, lateral and transverse diffusion, membrane asymmetry, and hydropathy plots. Describe the evidence for the lateral diffusion of membrane lipids and proteins. Contrast the rates for lateral diffusion with those for transverse diffusion.
20. Know about the functions of membranes; recognize correct statements about channel proteins, pumps, peripheral and integral membrane proteins.
21. Define and recognize examples of the following terms with words, equations, structures, examples, and/or diagrams. (Note that this list is NOT exhaustive)
    aldose, ketose, gel filtration
    alpha helix, glycoside
    amylopectin, hemiacetal
    anomer, Henderson-Hasselbalch eqn.
    beta sheet, parallel & antiparallel, hydrogen bond
    Bronsted acid & base, hydrophobic interaction
    buffering capacity, lipid bilayer
    cellulose, micelles
    chiral center, mutarotation
    chromatography, phospholipid
    dialysis, pyranose
    electrophoresis, reducing sugar
    enantiomer, salt bond or electrostatic interaction
    fatty acid, zwitterion
    furanose, primary, secondary, tertiary, quaternary, structures of proteins
    Edman degradation, 2-dimensional chromatography

Objectives for Unit 2

Enzymes

1. Describe behavior and properties of enzymes as catalysts; discuss theories of enzyme action, list ways in which catalytic activity may be regulated.
2. Understand the concept of free energy, be able to work problems involving DG, DG¡, and Keq (know and understand the equations).
3. Understand the derivation of the Michaelis-Menten (MM) equation and know all assumptions and conditions ( initial conditions, steady state, [S]>>>[E]); cast the MM equation into both v as a function of [S] and in the Lineweaver Burke form (double reciprocal plot); evaluate Km and Vm from plots and from equation form. Use both plots and the MM equation to evaluate the relationship of variables and constants in the MM equation. Calculate specific activities and turnover numbers of enzymes, as in HMWK problems.
4. Distinguish competitive, and non-competitive inhibition both graphically and mechanistically; know the uses for irreversible inhibitors.
5. Understand the basics of enzyme kinetics and reaction rates, transition states and the free energy of activation. Be able to work problems similar to those given in the homework. Understand the principle of transition state stabilization and its significance in enzyme catalysis; understand the origin of pH-activity relationships in enzyme catalyzed reactions.
6. Be prepared to answer questions concerned with lysozyme structure and lysozyme catalyzed reaction, the mechanism for catalysis, and the experimental support for the proposed mechanism.
7. Be prepared to answer questions concerned with chymotrypsin structure and serine protease catalyzed reactions, the experimental strategies used for studies of catalysis, covalent catalysis and chymotrypsin's catalytic triad. Understand the difference in specificity between chymotrypsin, trypsin and elastase. Know about examples of mutagenesis studies aimed at altering the specificity of chymotrypsin. Know that subtilisin contains a very similar active site, yet has no amino acid similarity to the chymotrypsin family (convergent evolution).
8. Discuss how allosteric enzymes differ from Michaelis-Menten enzymes; know about the concerted and sequential models of allosteric control . Know about subunit structure and cooperativity of oxygen binding of hemoglobin, aspartate transcarbamoylase (ACTase) structure and the reaction catalyzed by ACTase, the principles involved in substrate and inhibitor binding, and the experimental support for allosteric control. Understand proteolytic cleavage as a control mechanism for enzyme activity in vivo.

Objectives for Unit 3

Energy and Metabolism

1. Be familiar with the structures of ATP, NAD+, NADP+, FAD, Coenzyme A.
2. Be able to do free energy calculations involving redox potentials. Understand the different mechanisms cells can use to drive unfavorable reactions including [ATP]/[ADP][Pi]. Know how to solve problems involving coupled reactions.
3. Know the steps, intermediates and enzymes of glycolysis; know which steps involve cofactors, oxidation/reduction reactions, and ATP hydrolysis or production. Know which enzymes are involved in glycolytic regulation and what the regulatory molecules are. Know the possible fates of the pyruvate formed from glycolysis including lactate, acetyl CoA, etc.
4. Be familiar with the names of enzymes and the different types (or class) of reactions they catalyze (e.g., dehydrogenase, kinase, aldolase, phosphatase, isomerase, mutase, etc.). Be able to recognize the oxidized and reduced forms of molecules. Be able to recognize examples of condensation, dehydration, decarboxylation, etc.
5. Know the steps and intermediates of the citric acid cycle and be familiar with the enzymes involved at each step. Know which steps require cofactors, generate CO2 or GTP, etc. Which intermediates can be used as precursors in biosynthesis? Know about the regulation of the TCA cycle.
6. Understand the asymmetry and stereospecificity of enzymatic reactions (e.g., citrate, NADH).
7. Understand the meaning of redox potential and its relation to free energy changes. Know the major (both mobile and non-mobile) components of the respiratory chain, the electron-carrying groups. Know the entry points of NADH and FADH2 in the respiratory chain and understand the electron shuttle mechanisms.
8. Know the chemiosmotic hypothesis and the evidence supporting the model. Understand the proposed models for coupling oxidation (electron transport) and phosphorylation via proton gradients (proton motive force).
9. Know the gluconeogenesis pathway, how it differs from glycolysis, how many ATP are consumed and at what steps, and its regulatory mechanism.
10. Be familiar with glycogen structure, synthesis, and degradation, and the enzymes responsible for synthesis and breakdown.. Understand about the various mechanisms involved in the regulation of glycogen metabolism, and about the tissue specific nature of this regulation. Understand the reciprocal regulation mechanisms of (1) glycogen synthesis and degradation and (2) glycolysis and gluconeogenesis. Understand the relationships among the different metabolic pathways we have studied involving glucose and their coordinated regulation.
11. Be familiar with the aspects of fatty acid metabolism (particularly beta-oxidation) that were covered in lecture.
12. Know the cellular locations of the metabolic pathways and enzymes we have studied. Understand the importance of prosthetic groups, cofactors, isozymes, and kinase/phosphatase enzyme regulation.

Objectives for Unit 4

Nucleic Acids and Molecular Biology

1. Understand the flow of genetic information in biological systems and the cellular locations of these events.
2. Be able to draw the structures of the bases (adenine, guanine, thymine, uracil, cytosine) and ribonucleotides, deoxyribonucleotides, mono-, di-, and tri-phosphate-nucleotides that contain these bases. Know which bases are purines and which are pyrimidines.
3. Know the nomenclature used to describe nucleic acids. Know which bases hydrogen-bond and be able to determine the base sequence of an RNA transcript knowing the sequence of the coding (sense) or non-coding (template) strand. Understand chain polarity and the convention for writing base sequences (always 5' to 3').
4. Understand the important features of DNA structure: right and left handed double-helices, antiparallel, complementary strands, major and minor grooves, base pairing, denaturation, A-DNA, B-DNA, and Z-DNA.
5. Know the different types of enzymes that operate on nucleic acids: exonuclease (5'-3' and 3'-5'), endonuclease, DNase, RNase, restriction enzymes, topoisomerase, ligase, helicase, DNA polymerase, RNA polymerase, primase (in primosome). Know the different enzymes and proteins involved in bacterial replication including the significance of Okazaki fragments, leading and lagging strands, DNA polymerase I and III, replication forks, template strand. Understand the significance of the editing function of DNA polymerase. Know that cells have many DNA repair systems.
6. Understand the significance of base sequence and how it determines protein structure and function. Understand the importance of specific types of sequences such as OriC, palindromes, repeats, operators, etc.
7. Understand some of the basic concepts of recombinant DNA technology. Know about the functions and uses of restiction endonucleases, understand about plasmids and phages as vectors, and about the construction and use of genomic and cDNA libraries. Understand what Southern and Northern blotting mean. Know the basis of DNA sequence determination by the chain termination (dideoxy, Sanger) method. Understand the basis for the polymerase chain reaction (PCR). Understand the terms chromosome walking, cDNA, expression library, site directed mutagenesis.
8. Know the subunit composition (and functions of the subunits) of RNA polymerase core enzyme and holoenzyme and understand their mode of action. Know the following terms and how they relate to transcription: promoter, up-stream and down-stream sequences, start and termination sites, rho protein, TATA box, enhancer sequences, splicing and RNA secondary structure.
9. Be familiar with the general structure of tRNAs and the roles of codons and anticodons in translating the genetic code. Understand the degeneracy of the genetic code and wobble. Know the start and stop codons.
10. Know the (sequential) events of and components (nucleic acids, enzymes, and proteins) involved in protein synthesis: aminoacyl-tRNA synthetases, ribosomes (including their composition), A and P sites. Know that initiation factors (IF1, etc.), elongation factors (EF-Tu), GTP, termination factors also take part in translation.
11. Understand post-transcriptional processing of RNA and the events involved. What is RNA splicing, introns and exons, intervening sequences, the 5' cap and poly(A) tail on eukaryotic mRNA?
12. Understand the role of operators and repressors in the control of gene expression and their location and properties. Using the lac operon as a model, be able to describe the release of catabolite repression and the process of induction that occurs when the bacterial cell culture medium is changed from glucose to lactose. Include the roles of CAP protein, cAMP, repressor protein, inducer, and RNA polymerase.

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