Biochemical Techniques
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Course Introduction
Biochemical Techniques is a course designed to introduce students to the fundamental methods and instrumentation used in modern biochemical research. The course covers principles and applications of techniques such as spectroscopy, chromatography, electrophoresis, centrifugation, and various types of blotting (e.g., Western, Southern, and Northern). Students will learn how these techniques are used for the qualitative and quantitative analysis of proteins, nucleic acids, and other biomolecules. Emphasis is placed on experimental design, data analysis, troubleshooting, and understanding the strengths and limitations of each technique. The course combines theoretical instruction with hands-on laboratory experience to equip students with practical skills essential for research in biochemistry and related fields.
Recommended Textbook
Fundamentals of Biochemistry Life at the Molecular Level 5th Edition by Donald Voet
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Chapter 1: Introduction to the Chemistry of Life
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Q1) Gibbs free energy is defined as G = ______.
A) phylogenetic
B) negative
C) H<sub>2</sub>O
D) H°
E) H TS
F) halobacteria
G) open
H) entropy
I) flux
J) polymers
K) G°´
L) thermophiles
M) nitrogen
N) positive
Answer: E
Q2) Explain in one or two sentences why scientists believe that all living organisms are related (that they have all evolved from a common ancestor)?
Answer: Biochemically,all living organisms are very similar.
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Chapter 2: Water
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Q1) Which of the following statements about water is incorrect?
A)It is a small,polar molecule with a low dielectric constant.
B)It has a marked dipole moment.
C)It is largely hydrogen bonded,although any single H-bond exists only for a very short period of time (~10<sup>-12</sup> s ).
D)Acid-base reactions are very fast due to the mobility of hydronium ions in water which is a consequence of the ability of individual protons to "jump" from one water molecule to another.
E)It has a bent geometry with each O H bond approximately 0.958 Å long and with an O H bond energy of approximately 460 kJ/mol.
Answer: A
Q2) The pH at the midpoint in the titration of an acid with a base is
A)equal to the pK of the corresponding acid.
B)equal to the pK of the corresponding base.
C)equal to 14 minus the pK of the corresponding acid.
D)equal to 14 plus the pK of the corresponding base.
E)none of the above
Answer: A
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Chapter 3: Nucleotides, nucleic Acids, and Genetic Information
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Q1) After hybridization,the fragment of interest can be detected by ______.
A) diploid
B) phosphodiester
C) probe
D) pyrimidine
E) introns
F) cloning
G) transformed
H) exons
I) ampicillin
J) autoradiography
K) chain-terminator
L) purine
M) ester
Answer: J
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Chapter 4: Amino Acids
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Sample Questions
Q1) Which of the following amino acids has a sulfur atom in its side chain?
A)Asn
B)Ser
C)Phe
D)Met
E)Tyr
Q2) What is the three-letter abbreviation for the peptide valylarginylisoleucine?
A)Val Arg Ile
B)Val Agn Leu
C)Vln Arg Ile
D)Val Agn Isl
E)Vln Arg Leu
Q3) The tripeptide alanyllysylaspartate contains four ionizable groups with pK<sub>a</sub>s of 2.0,3.9,9.9,and 10.5.Calculate the pI for this molecule.
A)3)0
B)6)0
C)6)2
D)10.2
E)none of the above
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Chapter 5: Proteins: Primary Structure
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Q1) If the cDNA for a protein has been cloned,it may be possible to obtain large quantities of the protein by _________________ in bacteria.
A) electrophoresis
B) hydrophobic interaction
C) enzyme-linked immunosorbent assay
D) three-dimensional shape
E) N-terminal amino acid
F) negative charge
G) nucleases
H) chromophore
I) foaming
J) high level expression
K) 2-mercaptoethanol
L) positive charge
M) cation exchange
N) pI
O) chymotrypsin
P) C-terminal amino acid
Q) Sodium dodecyl sulfate
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Chapter 6: Proteins: Three-Dimensional Structure
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Q1) Repeating values of \(\phi\) and \(\varPsi\) make up predictable orientations of amino acid with a chain,this predictable orientation forms___________.
A) tertiary structure
B) keratin
C) molecular chaperones
D) hydropathy
E) cis
F) trans
G) sterically forbidden conformation
H) regular secondary structure
I) collagen
J) peptide bond
K) ribonuclease A (RNase A)
L) alpha
Q2) Evolutionary processes have
A)increased the stability of 4° structures.
B)decreased the number of subunits.
C)increased similarity amount 1° structures.
D)enhanced efficient folding pathways.
E)all of the above
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Page 8
Chapter 7: Protein Function Part I: Myoglobin and Hemoglobin
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Q1) The rearrangement of T state hemoglobin to the R state
A)occurs in each protein subunit independently when its heme binds oxygen.
B)requires the binding of at least three oxygen molecules.
C)increases the ion pairing interactions of the C-terminal amino acids.
D)involves the movement of the Fe(II)into the heme plane.
E)opens a central cavity for BPG binding.
Q2) The reaction of carbonic anhydrase catalyzes
A)the formation of carbamates with the concomitant release of protons.
B)the hydration of bicarbonate,resulting in the formation of carbonic acid.
C)the reduction of carbon dioxide with the concomitant consumption of protons.
D)the hydration of carbon dioxide,forming bicarbonate and protons.
E)the hydrolysis of carbamates with the concomitant consumption of protons.
Q3) Hemoglobin S,the variant responsible for the misshapen red blood cells characteristic of the disease sickle-cell anemia,is potentially advantageous to heterozygotes because it confers some level of resistance to the disease _________.
A)rickets
B)AIDS
C)cyanosis
D)polycythemia
E)malaria
Page 9
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Chapter 8: Carbohydrates
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Q1) Glucose,with its four chiral carbon atoms,has ______ possible stereoisomers.
A) eight
B) deoxy
C) peptidoglycan
D) axial
E) anomers
F) two
G) lactose
H) glyceraldehyde
I) amylopectin
J) epimers
K) sixteen
L) glycogen
M) equatorial
N) chitin
O) hyaluronic acid
P) amylose
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Chapter 9: Lipids and Biological Membranes
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Q1) Which of the following lipid types is the least polar?
A)triacylglycerols
B)glycerophospholipids
C)sphingolipids
D)gangliosides
E)glycolipids
Q2) A researcher is developing a new drug for the treatment of influenza.Based on the information presented in the text about the influenza virus,which of the following drug reactions describes the action of this new drug?
A)A drug which specifically interrupts the conformational change that results from the pH change in the endosome.
B)A drug which enhances the function of a normally quiescent enzyme responsible for cleavage of the carbohydrate component on glycophorin A proteins.
C)A drug which readily binds the hemagglutinin tag and results in conformational change.
D)All of the above are potential actions of this new drug.
E)None of the above is a plausible reaction for this new drug.
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Chapter 10: Membrane Transport
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Q1) In the intestinal epithelium,glucose enters the cell in symport with ______,which is subsequently ejected by ________.
A)K<sup>+</sup>;the Na<sup>+</sup>/K<sup>+</sup> pump
B)H<sup>+</sup>;oxidative metabolism
C)Na<sup>+</sup>;the Na<sup>+</sup>/K<sup>+</sup> pump
D)Na<sup>+</sup>;the GLUT1 transporter
E)Cl<sup>-</sup>;MDR transporter
Q2) Which of the following statements is true regarding ion channels?
A)Protein structure studies indicate a flexible pore size capable of transporting ions of varying size.
B)Ions require waters of hydration for sufficient passage through the entire channel.
C)Ion channels maintain sodium and potassium ion concentrations that are greater in the extracellular fluid than in the cytosol.
D)Channel rigidity minimizes energy states for appropriately sized ions.
E)None of the above is true regarding ion channels.
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Chapter 11: Enzyme Catalysis
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Q1) Which of the following is TRUE regarding cofactors?
A)Coenzymes are often separate from the enzyme and do not need recharged.
B)Metal ions must be covalently attached to function as a cofactor.
C)Cofactor is a broad term used for all enzyme "helpers".
D)Prosthetic groups can dissociate readily and be regenerated for use in another enzyme.
E)An apoenzyme implies that a cofactor is present.
Q2) If you add enzyme to a solution containing only the product(s)of a reaction,would you expect any substrate to form?
A)It depends on the time interval and temperature of reaction.
B)It depends on the concentration of products added.
C)It depends on the energy difference between E + P and the transition state.
D)All of the above may determine if product forms.
E)None of the above determines if product forms.
Q3) Proximity effects
A)result from active site specificity.
B)result from substrate channeling.
C)result in increased effective concentration of substrate.
D)lower the energy of activation.
E)All of the above of the above are correct.
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Chapter 12: Enzyme Kinetics, inhibition and Control
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Q1) This diagram refers to a (an)
A)Ping Pong reaction.
B)ordered bisubstrate reaction.
C)random bisubstrate reaction.
D)double order ping pong reaction
E)X,Y,and Z must be provided in order to answer correctly
Q2) A Lineweaver-Burk plot is also referred to as (I)a sigmoidal plot.
(II)a linear plot.
(III)a Michaelis-Menten plot.
(IV)a double reciprocal plot.
A)II
B)II,III
C)IV
D)II,IV
E)III,IV
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Chapter 13: Biochemical Signaling
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Q1) Sildenafil (Viagra®)
A)inhibits activity of a phosphoprotein phosphatase.
B)increases the relative concentration of cGMP.
C)increases the relative concentration of nitric oxide in the blood.
D)A,B,and C
E)A and B
Q2) Which of the following activities would lead to a decrease in the level of IP<sub>3</sub> in the cytosol?
A)increased activity of inositol polyphosphate 5-phosphatase
B)increased activity of phospholipase C
C)increased activation of protein kinase C
D)increased Ca<sup>2+</sup><sup> </sup>concentration
E)all of the above
Q3) Which of the following is always true of hormones?
A)Hormones always bind membrane bound receptors.
B)Hormones elicit specific cellular responses.
C)Hormones are secreted from a gland via a robust network of ducts.
D)The release of hormones is influenced by adrenal receptors.
E)All of the above are true.
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Chapter 14: Introduction to Metabolism
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Sample Questions
Q1) Consider the following metabolic reaction:
3-Phosphoglycerate \(\to\)2-Phosphoglycerate \(\Delta\)G°' = +4.40 kJ/mol
What is the \(\Delta\)G for this reaction when the concentration of 2-phosphoglycerate is 0.290 mM and the concentration of 3-phosphoglycerate is 2.90 mM at 37°C?
A)+10.3 kJ/mol
B)-1.53 kJ/mol
C)-1.30 kJ/mol
D)-5.93 kJ/mol
E)-4.40 kJ/mol
Q2) The conversion of triacylglycerides into fatty acids for the purpose of energy generation is an example of which of the following?
A)heterotropism
B)anaerobism
C)catabolism
D)anabolism
E)glycolysis
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Chapter 15: Glucose Catabolism
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Q1) Glucose is converted to ______ in skeletal muscle under anaerobic conditions.
A)lactate
B)acetyaldehyde
C)fructose
D)glycogen
E)acetyl-CoA
Q2) The reaction catalyzed by the enzyme aldolase has a \(\Delta\)G' +23 kJ/mol.In muscle cells,the reaction proceeds in this same,forward direction.How can this occur?
A)This \(\Delta\)G' means it is thermodynamically favored.
B)The enzyme changes the G of the reaction in cells to something favorable.
C)The concentration of reactant(s)must be significantly greater than product(s)in cells.
D)The concentration of product(s)must be significantly greater than reactant(s)in cells.
E)none of the above
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Chapter 16: Glycogen Metabolism and Gluconeogenesis
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Q1) Which of the following will NOT activate glycogen phosphorylase in the liver?
A)epinephrine
B)glucagon
C)eating a high carbohydrate meal
D)cAMP
E)All of the above will activate glycogen phosphorylase.
Q2) The breakdown of glycogen produces G1P.Which of the following is FALSE regarding G1P?
A)It is involved in the utilization of fructose by glycolysis in the liver.
B)It is involved in the utilization of fructose by glycolysis in the muscle.
C)It is involved in utilization of galactose by glycolysis.
D)It is converted to G6P by phosphoglucoisomerase (PGI).
E)It is produced as an intermediate in the central glycolytic sequence.
Q3) Phosphoprotein phosphatase -1 (PP1)
A)catalyzes the phosphorylation of phosphorylase kinase.
B)increases glycogen breakdown when active.
C)catalyzes the dephosphorylation of phosphorylase kinase.
D)catalyzes the dephosphorylation of glycogen phosphorylase.
E)Both C and Dare correct.
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Chapter 17: Citric Acid Cycle
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Q1) The citric acid cycle is also called the ______ cycle.
A) oxaloacetate
B) malate synthase
C) malate
D) malonate
E) Krebs
F) aconitase
G) succinyl-CoA synthetase
H) -ketoglutarate
I) carbon dioxide
J) malate dehydrogenase
K) isocitrate
L) succinate dehydrogenase
Q2) How many electrons are transferred from one acetyl group during the citric acid cycle?
A)2
B)4
C)6
D)8
E)10
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Page 19
Chapter 18: Electron Transport and Oxidative
Phosphorylation
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Q1) Consider the following half-reactions: \(\varepsilon\)<sup>o</sup>' = -0.315 V NAD<sup>+</sup> + H<sup>+</sup> + 2 e<sup>-</sup> \(\leftrightarrow\) NADH \(\varepsilon\)<sup>o</sup>' = 0.815 V 1/2 O<sub>2</sub> + 2 H<sup>+ </sup>+ 2 e<sup>- </sup>\(\leftrightarrow\) H<sub>2</sub>O What is the \(\varepsilon\)<sup>o</sup>'for the following redox equation? O<sub>2</sub> + 2 NADH + 2 H<sup>+ </sup> \(\to\) 2 H<sub>2</sub>O + 2 NAD<sup>+</sup>
A)-1.130 V
B)-0.500 V
C)+0.500 V
D)+1.130 V
E)none of the above
Q2) The structure of ATP synthase changes in conformation as a result of
A)the rotation of the and subunits.
B)the rotation of the subunit.
C)the rotation of ATP.
D)the rotation of the \(\gamma\) subunit.
E)all of the above.
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Chapter 19: Photosynthesis
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Q1) The major sugar transported to nonphotosynthesizing cells is
A)sucrose.
B)fructose.
C)UDP-glucose.
D)fructose-6-phosphate.
E)glyceraldehyde-3-phosphate.
Q2) In photorespiration,reduction of hydroxypyruvate to glycerate requires ____ and occurs in the ____.
A)NADPH;mitochondrion
B)NADH;peroxisome
C)NADPH;chloroplast
D)NADH;chloroplast
E)none of the above
Q3) In the dark reactions of photosynthesis,glyceraldehyde-3-phosphate is converted to dihydroxyacetone phosphate by___.
A)phosphoglucoisomerase
B)triose phosphate isomerase
C)glyceraldehyde-3-phosphate dehydrogenase
D)phosphotriose epimerase
E)phosphoglycerate kinase
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Chapter 20: Lipid Metabolism
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Q1) The activity of lipases is controlled in part by a process known as ____ which functions effectively due to the ____ where lipases are active.
A)interfacial activation;lipid-water interface
B)lipase activation;lipid-water interface
C)lipase activation;bile salt-water interface
D)micelle activation;water-salt interface
E)none of the above
Q2) Which of the following could be produced by the reaction of two or more of phosphatidic acids with two or more glycerol-3-phosphates?
(I)phosphatidylglycerol
(II)cardiolipin
(III)phosphatidylinositol
(IV)gangliosides
A)I,II,III,IV
B)II,III
C)I,III
D)I,II
E)III only
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22
Chapter 21: Amino Acid Metabolism
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Q1) The activity of bacterial glutamine synthetase is (I)inhibited allosterically by tryptophan. (II)inhibited allosterically by glucosamine-6-phosphate. (III)activated by nine allosteric feedback inhibitors. (IV)altered by covalent modification.
A)I,III
B)III,IV
C)I,II
D)I,IV
E)I,II,III,IV
Q2) Ketogenic amino acids can be converted to (I)glucose
(II)fatty acids
(III)ketone bodies (IV)pyruvate
A)I,III
B)II,IV
C)I,II
D)I,II,III
E)I,IV
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Page 23
Chapter 22: Mammalian Fuel Metabolism: Integration and Regulation
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Q1) Which of the following statements concerning the response of hepatocytes to insulin is FALSE?
A)Phosphorylase kinase is inactivated,decreasing glycogenolysis.
B)Glycogen synthase is activated.
C)Transcription of glucokinase increases.
D)Glucose uptake is increased in the liver.
E)The liver stores glucose as glycogen or triacylglycerols.
Q2) Synthesis of triacylglycerols is dependent on
A)the availability of glycerol-3-phosphate.
B)the uptake of glucose from the bloodstream.
C)the enzyme responsible for reduction of dihydroxyacetone phosphate.
D)the circulation of lipoprotein.
E)all of the above
Q3) Which of the following is part of the regulation of metabolism?
A)receptor tyrosine kinase
B)adenylate cyclase
C)phosphoinositide pathway
D)all of the above
E)none of the above
Page 24
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Chapter 23: Nucleotide Metabolism
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Q1) The end products of pyrimidine catabolism are
A)"uric acid and glyoxylic acid."
B)"uric acid and allantoin."
C)"\(\beta\)-alanineand \(\beta\)-aminoisobutyrate."
D)"\(\beta\)-alanine and \(\alpha\)-ketoglutarate."
E)"none of the above"
Q2) The synthesis of 5-phosphoribosylamine from PRPP is (I)inhibited allosterically by ATP and GTP. (II)activated by PRPP via feed forward activation. (III)controlled by cAMP dependent phosphorylation.
A)I,II,III,
B)I,II
C)II,III
D)II only
E)III only
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Chapter 24: Nucleic Acid Structure
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Q1) Indirect readout refers to (I)a protein's ability to form hydrogen bonds through a network of water molecules. (II)the 5' to 3' sequence on the opposite strand of a DNA helix. (III)a protein's ability to recognize a specific DNA sequence by its backbone conformation and flexibility.
(IV)the protein's ability to make hydrogen bonds with nonesterified phosphate oxygens
A)I,III,IV
B)II,IV
C)III,IV
D)I,III
E)I,IV
Q2) Hammerhead ribozyme
A)contains a catalytic core with a Fe<sup>2+</sup> at its center.
B)utilizes hydrogen bonding to position itself favorably for nucleophilic attack. C)enhances the rate of cleavage of itself by a factor of five.
D)A and C
E)B and C
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Chapter 25: DNA Replication, repair and Recombination
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Q1) Which of the following are TRUE about transposons?
(I)They are normal components of plasmids. (II)They often carry genes for antibiotic resistance. (III)They insert at a target sequence that is a repetitive unit of about 200 bp.
A)I,II,III
B)I,II
C)II,III
D)II only
E)III only
Q2) Which of the following describes the usefulness of primase?
A)It removes RNA bases and replaces them with DNA bases.
B)It seals nicks.
C)It adds an RNA starter sequence which can be elongated.
D)It is only required on the leading strand.
E)It unwinds the DNA.
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Chapter 26: Transcription and Rna Processing
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Q1) Which of the following best describes the cap nucleotide on a eukaryotic mRNA?
A)A
B)C
C)m<sup>7</sup>G
D)T
E)U
Q2) In which of the following types of eukaryotics RNAs are introns completely removed most of the time,although exceptions can occur?
(I)mRNA
(II)rRNA
(III)tRNA
A)I,II
B)I,III
C)II,III
D)I,II,III
E)none of the above
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Chapter 27: Protein Synthesis
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Q1) The Shine-Dalgarno sequence is
A)used to align ribosome subunits and initiation translation.
B)functions as a stop codon.
C)is used to align RNA polymerase.
D)is used during transcription termination.
E)all of the above
Q2) The structural similarities of EF-G and the EF-Tu·tRNA complex indicate
A)EF-G may be used as a substitute for EF-Tu.
B)EF-G may play a role in displacing the tRNA growing peptide.
C)EF-G may mediate a change in conformation that facilitates EF-Tu binding.
D)EF-G may be involved in facilitating binding of initiation factors.
E)EF-G and EF-Tu are both required for proper initiation.
Q3) Codons that do not specify amino acids but signal the ribosome to terminate polypeptide chain elongation are termed
A)frameshifts.
B)anticodon.
C)Stop codons.
D)suppressors.
E)terminators.
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Page 29
Chapter 28: Regulation of Gene Expression
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Q1) In eukaryotic cells,methylated bases are present in palindromic ______,which are often found in upstream promoter regions.
A)CpG islands
B)ORFs
C)SNPs
D)LINEs
E)SINEs
Q2) A database of _____ is being compiled to examine possible genetic links to the susceptibility to disease or infection..
A)CpG islands
B)ORFs
C)SNPs
D)LINEs
E)SINEs
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