Perspectives of enzyme engineering MCQs With Answer

Perspectives of Enzyme Engineering MCQs With Answer

This quiz collection provides M.Pharm students with focused multiple-choice questions on enzyme engineering—an essential area for modern pharmaceutical bioprocessing and drug synthesis. It covers core concepts such as directed evolution, rational and semi‑rational design, computational protein design, immobilization strategies, cofactor engineering, kinetics, stability improvements, and applications in chiral drug synthesis and biocatalysis. Questions emphasize mechanistic understanding, experimental approaches (mutagenesis, screening, expression optimization), and practical considerations for industrial enzyme implementation. Use these MCQs to deepen conceptual knowledge, prepare for exams, and bridge academic theory with real‑world enzyme engineering used in pharmaceutical development and green chemistry.

Q1. Which parameter best describes catalytic efficiency of an enzyme under second‑order conditions and is often used to compare mutated variants?

• kcat
• Km
• kcat/Km
• Vmax

Correct Answer: kcat/Km

Q2. Directed evolution typically requires two iterative steps; which pair correctly describes these steps?

• Homologous recombination and X‑ray crystallography
• Random or targeted mutagenesis and high‑throughput screening/selection
• Site‑directed mutagenesis and circular dichroism
• Protein glycosylation and surface display purification

Correct Answer: Random or targeted mutagenesis and high‑throughput screening/selection

Q3. Which technique is most suited for introducing a high frequency of point mutations across an entire gene to generate diversity for evolution experiments?

• Site‑directed mutagenesis
• Error‑prone PCR
• Affinity chromatography
• Mass spectrometry

Correct Answer: Error‑prone PCR

Q4. Rational design in enzyme engineering primarily relies on which resource to guide specific amino acid substitutions?

• Random library creation without structural data
• High‑resolution structural data and mechanistic understanding
• Purely empirical fermentation conditions
• Patent literature only

Correct Answer: High‑resolution structural data and mechanistic understanding

Q5. Semi‑rational or focused library design aims to balance between random mutagenesis and rational design by:

• Mutating every residue in the protein sequentially
• Targeting residues informed by structure or sequence conservation to create smaller, informed libraries
• Avoiding any structural information and using only consensus sequences
• Only altering codon usage without changing amino acids

Correct Answer: Targeting residues informed by structure or sequence conservation to create smaller, informed libraries

Q6. Which computational tool or approach is frequently used to predict stabilizing mutations and redesign active sites in modern enzyme engineering?

• Rosetta protein design and molecular dynamics simulations
• Thin layer chromatography
• SDS‑PAGE electrophoresis
• Gas chromatography with flame ionization detector

Correct Answer: Rosetta protein design and molecular dynamics simulations

Q7. Immobilization of enzymes on solid supports commonly provides which advantage for industrial biocatalysis?

• Reduced substrate specificity
• Improved operational stability and easy enzyme reuse
• Complete loss of activity at ambient temperature
• Higher Km values only

Correct Answer: Improved operational stability and easy enzyme reuse

Q8. Which experimental strategy is most appropriate to evolve an enzyme for improved thermostability?

• Selection at progressively higher temperatures combined with screening for retained activity
• Reducing pH during selection only
• Removing disulfide bonds systematically
• Overexpressing the enzyme without selection

Correct Answer: Selection at progressively higher temperatures combined with screening for retained activity

Q9. The catalytic triad in serine proteases typically consists of which combination of amino acids?

• Serine, Histidine, Aspartate
• Lysine, Proline, Glycine
• Cysteine, Tyrosine, Glutamate
• Alanine, Valine, Leucine

Correct Answer: Serine, Histidine, Aspartate

Q10. In cofactor engineering for oxidoreductases, a common industrial aim is to:

• Replace a catalytic residue with zinc
• Alter enzyme preference from NADPH to NADH to lower process cost
• Prevent any cofactor binding to increase substrate range
• Eliminate all redox activity

Correct Answer: Alter enzyme preference from NADPH to NADH to lower process cost

Q11. Which metric must be considered when engineering an enzyme for enhanced enantioselectivity in chiral drug synthesis?

• Enantiomeric excess (ee) and enantioselectivity (E) value
• Only molecular weight of enzyme
• Length of expression vector
• pKa of unrelated buffer species

Correct Answer: Enantiomeric excess (ee) and enantioselectivity (E) value

Q12. Ancestral sequence reconstruction is used in enzyme engineering primarily to:

• Create synthetic cofactors
• Infer ancient protein sequences that are often more stable and promiscuous
• Delete regulatory elements in plasmids
• Measure Km values directly

Correct Answer: Infer ancient protein sequences that are often more stable and promiscuous

Q13. Which of the following is NOT a typical mechanism by which mutations increase enzyme thermostability?

• Introduction of additional salt bridges and hydrogen bonds
• Increasing internal packing and hydrophobic interactions
• Increasing surface loop flexibility to promote unfolding
• Introduction of disulfide bonds where structurally tolerated

Correct Answer: Increasing surface loop flexibility to promote unfolding

Q14. Cross‑linked enzyme aggregates (CLEAs) are an immobilization method; what is a primary benefit of CLEAs?

• They require fusion to large tags for activity
• They form carrier‑free, highly concentrated biocatalysts with high stability
• They always decrease enzyme activity by 90%
• They make enzymes water‑insoluble by glycosylation only

Correct Answer: They form carrier‑free, highly concentrated biocatalysts with high stability

Q15. Which assumption is inherent in standard Michaelis–Menten kinetics?

• Product formation is instantaneous and irreversible
• Substrate concentration is much greater than enzyme concentration and steady‑state is achieved for the enzyme–substrate complex
• Enzyme concentration is higher than substrate concentration
• Km is always equal to Kd

Correct Answer: Substrate concentration is much greater than enzyme concentration and steady‑state is achieved for the enzyme–substrate complex

Q16. Site‑saturation mutagenesis at a catalytic residue position is most useful when the goal is to:

• Completely delete the gene
• Explore all 20 amino acid substitutions at that specific position to tune activity or specificity
• Measure protein concentration by Bradford assay
• Only change codon usage without altering amino acid

Correct Answer: Explore all 20 amino acid substitutions at that specific position to tune activity or specificity

Q17. Which strategy helps maintain enzyme activity in nonaqueous or low‑water organic media commonly used for synthetic chemistry?

• Using only soluble enzymes with no stabilizing agents
• Lyophilization, immobilization, and controlling microenvironment water activity
• Adding proteases to degrade the enzyme
• Using very high temperatures without stabilization

Correct Answer: Lyophilization, immobilization, and controlling microenvironment water activity

Q18. In multi‑enzyme cascade reactions for API synthesis, substrate channeling is beneficial because it:

• Increases diffusion distances between enzymes
• Reduces intermediate accumulation and increases overall pathway flux
• Always requires membrane compartments
• Prevents cofactor recycling

Correct Answer: Reduces intermediate accumulation and increases overall pathway flux

Q19. When expressing engineered enzymes in E. coli, codon optimization and co‑expression of chaperones are used to:

• Reduce protein solubility intentionally
• Improve soluble expression and correct folding to obtain active enzyme
• Introduce glycosylation identical to eukaryotes
• Remove all disulfide bonds automatically

Correct Answer: Improve soluble expression and correct folding to obtain active enzyme

Q20. Regulatory considerations for using engineered enzymes as therapeutic biologics include which of the following concerns?

• Immunogenicity, product consistency, and demonstration of safety and efficacy
• Only room temperature storage without documentation
• Irrelevance of manufacturing controls for biologics
• Using any sequence regardless of similarity to human proteins is automatically acceptable

Correct Answer: Immunogenicity, product consistency, and demonstration of safety and efficacy

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