MChem Master of Chemistry Biochemistry and Chemical Biology 2 — Questions and Answers
Question 1: What distinguishes click chemistry reactions, such as the Cu-catalyzed azide-alkyne cycloaddition (CuAAC)?
- They require harsh acidic or basic conditions
- They are modular, high-yielding, and produce only innocuous byproducts under mild conditions (Correct answer)
- They exclusively form C–C bonds
- They require protecting groups and multiple purification steps
Correct answer: They are modular, high-yielding, and produce only innocuous byproducts under mild conditions
Click reactions are defined by their high efficiency, selectivity, and bioorthogonality, making them ideal for labeling biomolecules in living cells.
Question 2: What is the principle behind FRET (Förster Resonance Energy Transfer) in biochemical assays?
- Electrons transfer directly between two fluorophores via tunneling
- Non-radiative energy transfers from a donor fluorophore to an acceptor when they are within ~10 nm of each other (Correct answer)
- Two fluorophores absorb the same photon simultaneously
- A fluorophore transfers energy to oxygen, generating reactive singlet oxygen
Correct answer: Non-radiative energy transfers from a donor fluorophore to an acceptor when they are within ~10 nm of each other
FRET occurs via dipole-dipole coupling between a donor and acceptor fluorophore; its efficiency falls off as the inverse 6th power of distance, making it a molecular ruler for ~1–10 nm distances.
Question 3: In proteomics, what is the purpose of tryptic digestion before mass spectrometry analysis?
- To denature proteins for better ionization
- To cleave proteins into peptides at Arg and Lys residues, generating a reproducible, manageable set of fragments (Correct answer)
- To remove glycosylation from proteins before analysis
- To cross-link proteins to preserve their native state
Correct answer: To cleave proteins into peptides at Arg and Lys residues, generating a reproducible, manageable set of fragments
Trypsin cleaves specifically after Arg and Lys residues (except before Pro), generating peptides in the 500–3000 Da range optimal for LC-MS/MS identification.
Question 4: What is the chemical basis of disulfide bond formation in protein folding?
- Two histidine residues coordinate a zinc ion to form a structural crosslink
- Two cysteine thiol groups (-SH) are oxidized to form a covalent S–S bridge (Correct answer)
- A glutamine and asparagine residue condense to form an isopeptide bond
- Serine and threonine hydroxyl groups condense to form ether linkages
Correct answer: Two cysteine thiol groups (-SH) are oxidized to form a covalent S–S bridge
Disulfide bonds form when two cysteine thiol groups lose electrons (oxidation) in the oxidizing environment of the ER lumen, catalyzed by protein disulfide isomerase (PDI).
Question 5: What is the function of ubiquitin in protein degradation?
- It directly unfolds proteins for degradation by the proteasome
- It is conjugated to target proteins as a polyubiquitin tag that directs them to the 26S proteasome (Correct answer)
- It serves as a cofactor that activates the proteasome's proteolytic subunits
- It prevents protein aggregation by acting as a molecular chaperone
Correct answer: It is conjugated to target proteins as a polyubiquitin tag that directs them to the 26S proteasome
A cascade of E1 (activating), E2 (conjugating), and E3 (ligase) enzymes attach polyubiquitin chains (typically Lys48-linked) to substrates, marking them for 26S proteasomal degradation.
Question 6: What chemical feature distinguishes RNA from DNA that makes RNA more susceptible to alkaline hydrolysis?
- RNA has uracil instead of thymine, which is less stable
- The 2'-hydroxyl group of ribose acts as an intramolecular nucleophile to cleave the phosphodiester backbone (Correct answer)
- RNA has a single strand that is less protected from hydrolysis
- RNA lacks the methyl group on pyrimidines that stabilizes DNA
Correct answer: The 2'-hydroxyl group of ribose acts as an intramolecular nucleophile to cleave the phosphodiester backbone
In RNA, the 2'-OH attacks the adjacent phosphodiester bond in a transesterification reaction, forming a 2',3'-cyclic phosphate intermediate and cleaving the backbone.
MChem Master of Chemistry Biochemistry and Chemical Biology 2
What distinguishes click chemistry reactions, such as the Cu-catalyzed azide-alkyne cycloaddition (CuAAC)?