Production of regulatory proteins MCQs With Answer

Introduction:

This collection of MCQs on the production of regulatory proteins is tailored for M. Pharm students studying Advanced Pharmaceutical Biotechnology. The quiz focuses on upstream and downstream strategies, expression systems, folding and secretion, post‑translational modifications, purification tactics, analytical assays and regulatory expectations. Questions probe practical decision‑making — for example, choosing expression hosts for complex glycoproteins, preventing inclusion body formation, optimizing glycosylation, and ensuring product quality and safety under cGMP. These items are intended to deepen understanding, prepare for exams and encourage application of theoretical concepts to real manufacturing and quality problems encountered when producing therapeutic regulatory proteins.

Q1. Which promoter is most commonly used for very high-level recombinant protein expression in E. coli when using T7 RNA polymerase-based systems?

• lac promoter
• araBAD promoter
• T7 promoter
• trp promoter

Correct Answer: T7 promoter

Q2. Which signal peptide is frequently used to direct recombinant regulatory proteins to the periplasm or secretion pathway in E. coli expression systems?

• PelB leader sequence
• Signal peptide from human insulin
• Kozak sequence
• SV40 nuclear localization signal

Correct Answer: PelB leader sequence

Q3. Codon optimization of a gene for expression in a chosen host primarily improves which aspect of production?

• Promoter strength
• mRNA splicing efficiency
• Translation efficiency by matching host codon usage
• Secretion efficiency through signal peptides

Correct Answer: Translation efficiency by matching host codon usage

Q4. Formation of inclusion bodies during high-level expression in E. coli is most often a consequence of which factor?

• Excessive post-translational glycosylation
• Protease-mediated degradation
• Rapid overexpression causing misfolding and aggregation
• Inadequate transcriptional initiation

Correct Answer: Rapid overexpression causing misfolding and aggregation

Q5. For producing regulatory proteins that require complex, human-like N-linked glycosylation, which expression host is preferred?

• Escherichia coli
• Bakery yeast (S. cerevisiae) without engineering
• Mammalian cell lines such as CHO cells
• Standard Baculovirus-insect cell systems without modification

Correct Answer: Mammalian cell lines such as CHO cells

Q6. Which molecular chaperone system is commonly co-expressed in E. coli to assist folding of recombinant regulatory proteins and reduce aggregation?

• Clp protease complex
• GroEL/GroES chaperonin system
• Sec translocon
• Ribosomal rescue factor

Correct Answer: GroEL/GroES chaperonin system

Q7. A functional Sec-dependent signal peptide for secretion typically contains which combination of features?

• A hydrophobic core with an N-terminal positively charged region and a signal peptidase cleavage site
• A C-terminal KDEL retention motif
• A proline-rich transmembrane helix with no cleavage site
• An internal glycosylation site and zinc-finger motif

Correct Answer: A hydrophobic core with an N-terminal positively charged region and a signal peptidase cleavage site

Q8. Which site-specific protease is widely used to cleave fusion tags from recombinant regulatory proteins because of its high sequence specificity and minimal off-target cleavage?

• Thrombin
• Enterokinase
• TEV protease
• Trypsin

Correct Answer: TEV protease

Q9. To humanize glycan structures on a recombinant protein produced in CHO cells, which strategy is most appropriate?

• Lowering culture temperature during production
• Using codon optimization of the gene
• Glycoengineering the host cell line to modify glycosyltransferase expression
• Adding detergents to the feed to alter glycosylation

Correct Answer: Glycoengineering the host cell line to modify glycosyltransferase expression

Q10. Which downstream purification method is particularly effective at removing endotoxin (lipopolysaccharide) from recombinant proteins produced in E. coli?

• Simple heat inactivation of the lysate
• Size-exclusion chromatography alone
• Ionic exchange chromatography optimized for endotoxin binding and removal
• Direct lyophilization of crude lysate

Correct Answer: Ionic exchange chromatography optimized for endotoxin binding and removal

Q11. Which upstream process modification commonly improves soluble yield and reduces aggregation of recombinant regulatory proteins in microbial expression systems?

• Increasing inducer concentration to maximum
• Lowering cultivation or induction temperature
• Raising dissolved oxygen setpoint to highest level
• Shortening induction time to minutes

Correct Answer: Lowering cultivation or induction temperature

Q12. When producing a secreted human cytokine that requires correct folding and disulfide bond formation, which eukaryotic host is most often chosen for commercial manufacturing?

• Escherichia coli cytosol
• Pichia pastoris without humanization
• Chinese Hamster Ovary (CHO) cells
• Unmodified Saccharomyces cerevisiae

Correct Answer: Chinese Hamster Ovary (CHO) cells

Q13. Which fusion tag is particularly effective at enhancing the solubility of difficult-to-fold recombinant regulatory proteins expressed in E. coli?

• Polyhistidine (His) tag
• Maltose-binding protein (MBP) tag
• FLAG epitope tag
• Short myc tag

Correct Answer: Maltose-binding protein (MBP) tag

Q14. For membrane-associated regulatory proteins, which class of detergent is generally preferred to solubilize protein while preserving native-like structure for downstream purification and characterization?

• Ionic detergents such as SDS
• Non-ionic or mild zwitterionic detergents such as DDM or CHAPS
• Strong denaturants like urea
• High concentration salts without detergents

Correct Answer: Non-ionic or mild zwitterionic detergents such as DDM or CHAPS

Q15. Which analytical approach is essential to confirm the functional activity of a recombinant regulatory protein such as a cytokine or growth factor?

• SDS-PAGE staining to check purity
• Mass spectrometry to confirm mass
• Cell-based bioassay measuring biological response
• UV absorbance at 280 nm for concentration

Correct Answer: Cell-based bioassay measuring biological response

Q16. Persistent proteolytic degradation of a recombinant regulatory protein during fermentation can be most directly mitigated by which strategy?

• Increasing aeration only
• Using protease-deficient host strains or protease knockouts
• Adding high concentrations of glucose at induction
• Using stronger promoters to outcompete degradation

Correct Answer: Using protease-deficient host strains or protease knockouts

Q17. In yeast expression systems, which leader sequence is commonly used to direct efficient secretion of recombinant human proteins into the media?

• PelB leader
• Alpha-mating factor prepro (α-factor) signal
• OmpA signal peptide
• Signal recognition particle (SRP) binding motif

Correct Answer: Alpha-mating factor prepro (α-factor) signal

Q18. In E. coli, formation of disulfide bonds required for many regulatory proteins is best supported in which cellular compartment by the Dsb pathway?

• Cytosol
• Periplasmic space
• Mitochondrial matrix
• Endoplasmic reticulum

Correct Answer: Periplasmic space

Q19. If a recombinant regulatory protein carries an N-terminal His6-tag, which purification method provides the most selective initial capture from clarified lysate?

• Hydrophobic interaction chromatography (HIC)
• Size-exclusion chromatography
• Immobilized metal affinity chromatography (IMAC) such as Ni-NTA
• Anion exchange at low salt

Correct Answer: Immobilized metal affinity chromatography (IMAC) such as Ni-NTA

Q20. Which regulatory requirement is absolutely central for the commercial manufacture of therapeutic regulatory proteins to ensure product quality, safety and traceability?

• Conducting only GLP non-clinical studies
• Adherence to current Good Manufacturing Practice (cGMP)
• Monitoring by local academic review boards only
• Using animal-derived media components exclusively

Correct Answer: Adherence to current Good Manufacturing Practice (cGMP)

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