Microbial biotransformation of steroids MCQs With Answer

Microbial biotransformation of steroids MCQs With Answer

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Introduction

This quiz collection on Microbial Biotransformation of Steroids is tailored for M.Pharm students preparing for advanced pharmaceutical biotechnology exams. It focuses on microbial pathways, key enzymes (e.g., cytochrome P450 monooxygenases, 3-ketosteroid dehydrogenases, 9α-hydroxylases), important bacterial and fungal biocatalysts (Mycobacterium, Rhodococcus, Corynebacterium, Cunninghamella, Aspergillus), and industrial conversions such as steroid side-chain cleavage and regio- and stereoselective hydroxylations. Questions probe mechanisms, cofactors, bioprocess strategies, genetic approaches to improve yields, and analytical considerations. The set is designed to deepen conceptual understanding and application of microbial transformations in steroid drug manufacture, quality control, and metabolic engineering.

Q1. Which enzyme class is most commonly responsible for regioselective hydroxylation (e.g., 11β- or 14α-hydroxylation) of steroid substrates in microbial biotransformations?

  • Flavin monooxygenases
  • Cytochrome P450 monooxygenases
  • Peroxidases
  • Hydrolases

Correct Answer: Cytochrome P450 monooxygenases

Q2. Which genus is most frequently exploited industrially for conversion of phytosterols to 17-ketosteroids (e.g., androstenedione) via side-chain degradation?

  • Rhodococcus
  • Mycobacterium
  • Cunninghamella
  • Aspergillus

Correct Answer: Mycobacterium

Q3. The enzyme 3-ketosteroid Δ1-dehydrogenase catalyzes which type of reaction in steroid metabolism?

  • Introduction of a double bond between C1 and C2 (Δ1-dehydrogenation)
  • Hydroxylation at C11
  • Cleavage of the side chain at C17
  • Reduction of a keto group to hydroxyl

Correct Answer: Introduction of a double bond between C1 and C2 (Δ1-dehydrogenation)

Q4. In microbial steroid biotransformation, which cofactor is most commonly required by dehydrogenases involved in oxidation-reduction of steroid keto/hydroxy groups?

  • FADH2
  • ATP
  • NAD+ / NADP+
  • CoA

Correct Answer: NAD+ / NADP+

Q5. 9α-hydroxylation of 3-ketosteroids in bacteria often initiates which downstream process in steroid catabolism?

  • Side-chain cleavage at C17
  • A-ring aromatization
  • A-ring cleavage leading to ring B–D opening and steroid nucleus degradation
  • C20–C21 epimerization

Correct Answer: A-ring cleavage leading to ring B–D opening and steroid nucleus degradation

Q6. Cunninghamella and Rhizopus are fungal biocatalysts commonly used for steroid transformations. Which transformation are these fungi especially known for?

  • Selective microbial glycosylation of steroids
  • Regio- and stereoselective hydroxylation (e.g., 11α, 6β)
  • Oxidative ring cleavage of the steroid nucleus
  • Large-scale side-chain removal to form androstenedione

Correct Answer: Regio- and stereoselective hydroxylation (e.g., 11α, 6β)

Q7. Which bioprocess strategy is most effective to reduce further degradation of a desired hydroxylated steroid product by the whole-cell catalyst?

  • Increasing inoculum density without substrate feeding control
  • Using wild-type strains with long incubation times
  • Knockout or downregulation of catabolic genes that attack the product
  • Raising temperature above optimum to denature degradative enzymes

Correct Answer: Knockout or downregulation of catabolic genes that attack the product

Q8. Which analytical technique is most appropriate for confirming positional and stereochemical hydroxylation on a steroid product?

  • Gas chromatography with flame ionization detector (GC-FID) alone
  • NMR spectroscopy
  • Infrared spectroscopy (IR) only
  • Colorimetric assay for total hydroxyl groups

Correct Answer: NMR spectroscopy

Q9. A common industrial route to cortisone-like compounds employed microbial 11β-hydroxylation. What is a major advantage of using microbial hydroxylation over classical chemical methods?

  • Microbial reactions are always faster than chemical synthesis
  • Microbial hydroxylation provides high regio- and stereoselectivity under mild conditions
  • Microbes do not require cofactors or oxygen for hydroxylation
  • Chemical methods cannot introduce hydroxyl groups at all

Correct Answer: Microbial hydroxylation provides high regio- and stereoselectivity under mild conditions

Q10. Which enzyme complex is commonly implicated in bacterial degradation of cholesterol and steroid side chains via multistep oxidation and β-oxidation-like reactions?

  • 3β-hydroxysteroid dehydrogenase only
  • Multienzyme complexes including acyl-CoA ligases, monooxygenases, dehydrogenases and thiolases
  • Single cytochrome c enzyme
  • Glycosyltransferases

Correct Answer: Multienzyme complexes including acyl-CoA ligases, monooxygenases, dehydrogenases and thiolases

Q11. In whole-cell steroid biotransformations, substrate solubility is often limiting. Which approach is commonly used to improve apparent solubility without denaturing cells?

  • Use of organic cosolvents at high concentration (>50%)
  • Emulsification with biocompatible surfactants or cyclodextrins
  • Heating to increase solubility beyond cell tolerance
  • Alkalinization to pH 12

Correct Answer: Emulsification with biocompatible surfactants or cyclodextrins

Q12. Which genetic engineering approach can increase steroid hydroxylation activity in a microbial host?

  • Deletion of the monooxygenase gene
  • Overexpression of the relevant cytochrome P450 and its redox partner
  • Knockout of all dehydrogenases
  • Overexpression of ribosomal proteins only

Correct Answer: Overexpression of the relevant cytochrome P450 and its redox partner

Q13. Stereospecific epimerization at C20 of corticosteroid substrates in microbial biotransformation is typically catalyzed by which enzyme type?

  • Epimerase or dehydrogenase/reductase pair operating via oxidation–reduction
  • Hydrolase acting on esters only
  • Lipase-like enzymes
  • Transaminases

Correct Answer: Epimerase or dehydrogenase/reductase pair operating via oxidation–reduction

Q14. Microbial biotransformation sometimes yields multiple regioisomers. Which process parameter most directly influences regioselectivity of hydroxylation?

  • The brand of culture flask used
  • Expression level and substrate-binding pocket architecture of the monooxygenase
  • Ambient humidity in the lab
  • Presence of exogenous proteases

Correct Answer: Expression level and substrate-binding pocket architecture of the monooxygenase

Q15. Which statement correctly contrasts isolated enzyme biocatalysis with whole-cell biotransformation for steroid conversions?

  • Isolated enzymes always require no cofactors; whole cells never need cofactors.
  • Isolated enzymes allow finer control of reaction conditions and cofactor recycling but may require enzyme stabilization; whole cells provide intrinsic cofactor regeneration and membrane transport systems.
  • Whole-cell systems always give higher purity products than isolated enzymes.
  • Isolated enzymes cannot catalyze hydroxylation reactions.

Correct Answer: Isolated enzymes allow finer control of reaction conditions and cofactor recycling but may require enzyme stabilization; whole cells provide intrinsic cofactor regeneration and membrane transport systems.

Q16. Which metabolic engineering modification can decrease degradation of an intermediate steroid product by a microbial host?

  • Overexpressing downstream catabolic enzymes
  • Deleting genes encoding enzymes that further metabolize the intermediate
  • Increasing intracellular protease expression
  • Adding random mutagens during production runs

Correct Answer: Deleting genes encoding enzymes that further metabolize the intermediate

Q17. For scalable production of a hydroxylated steroid, which downstream processing consideration is most critical immediately after the biotransformation to prevent product loss?

  • Rapid cell removal and quenching to stop enzymatic degradation
  • Storing the broth at room temperature for several days
  • Adding metal ions to the broth to catalyze further oxidation
  • Slow aeration to let microbes adapt

Correct Answer: Rapid cell removal and quenching to stop enzymatic degradation

Q18. Which enzyme family is known for catalyzing steroid A-ring isomerization (e.g., conversion of Δ5 to Δ4 steroids) during microbial transformations?

  • Ketoisomerases such as 3-ketosteroid isomerase (KSI)
  • Glycosidases
  • DNA polymerases
  • Phosphatases

Correct Answer: Ketoisomerases such as 3-ketosteroid isomerase (KSI)

Q19. Which factor most strongly affects the enantioselectivity of a microbial steroid reduction?

  • The water hardness in the laboratory
  • The stereochemistry of the enzyme active site and orientation of substrate binding
  • The color of the culture medium
  • The type of plasticware used to store substrate

Correct Answer: The stereochemistry of the enzyme active site and orientation of substrate binding

Q20. In screening microbial strains for a desired steroid transformation, which initial assay gives the most direct evidence of conversion and helps prioritize strains for characterization?

  • Visual inspection of culture color change only
  • Thin-layer chromatography (TLC) or HPLC analysis showing substrate disappearance and product formation
  • Measuring optical density (OD600) alone
  • Gram staining of the isolates

Correct Answer: Thin-layer chromatography (TLC) or HPLC analysis showing substrate disappearance and product formation

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