20. Although ribonucleoside triphosphates (rNTPs) are present at approximately IO-fold higher concentration than deoxyribo-nucleoside triphosphates (dNTPs) in the

cell but they are incorporated into DNA at a rate that is more than 1000-fold lower than dNTPs. This is because

(1) DNA polymerase cannot discriminate between dNTPs and rNTPs. But as soon as rNTPs areincorporated in the DNA chain, they are hydrolyzed due to the presence of 2′-OH group.

(2) DNA polymerase cannot dtscrirntnate between dNTPs and rNTPs. But IIS soon as rNTPs are incorporated in the DNA chain, they are excised by the proof reading activity of DNA polymerase.

(3) DNA polymerase efficiently discrtrrunatee. Between rNTPs and dNTPs, because its nucleotide binding pocket cannot accommodate a 2′-OH on the

incoming nucleotide.
(4) DNA polymerase cannot discriminate between rNTPs and dNTPs. Since the rate of transcription jncell is 10⁶ times faster than replication, it cannot compete with RNA polymerase for rNTPs.

Introduction

Inside cells, ribonucleoside triphosphates (rNTPs) are generally present at concentrations roughly 10 times higher than deoxyribonucleoside triphosphates (dNTPs). However, DNA polymerases incorporate rNTPs into DNA at rates more than 1000-fold lower than dNTPs. This remarkable selectivity is essential to maintain DNA integrity and prevent errors during replication. Understanding the molecular mechanism behind this discrimination is critical for insights into DNA replication fidelity.

Cellular Concentrations vs. Incorporation Rates

• rNTPs: Abundant substrates primarily used for RNA synthesis during transcription.

• dNTPs: Less abundant but the correct substrates for DNA synthesis during replication.

Despite the higher abundance of rNTPs, DNA polymerase preferentially incorporates dNTPs, ensuring DNA is composed of deoxyribonucleotides rather than ribonucleotides.

Molecular Basis of Discrimination by DNA Polymerase

Key Reason:
DNA polymerase’s nucleotide binding pocket is structurally designed to exclude the 2′-OH group present on ribonucleotides (rNTPs) but absent on deoxyribonucleotides (dNTPs).

• The presence of the 2′-OH group on rNTPs causes steric hindrance within the active site of DNA polymerase.

• This steric clash prevents efficient binding and incorporation of rNTPs.

• Consequently, DNA polymerase exhibits high fidelity and selectivity for dNTPs.

Evaluation of Options

1. DNA polymerase cannot discriminate but rNTPs are hydrolyzed after incorporation due to 2′-OH:

   • Incorrect. DNA polymerase discriminates before incorporation; rNTPs are rarely incorporated.

2. DNA polymerase cannot discriminate but proofreading excises rNTPs after incorporation:

   • Incorrect. Proofreading mainly removes mismatched bases, not ribonucleotides incorporated at very low frequency.

3. DNA polymerase efficiently discriminates because its nucleotide binding pocket cannot accommodate 2′-OH on rNTPs:

   • Correct. This structural discrimination is the primary reason for low rNTP incorporation.

4. DNA polymerase cannot discriminate and transcription outcompetes replication for rNTPs:

   • Incorrect and unrelated to polymerase substrate specificity.

Correct Answer

(3) DNA polymerase efficiently discriminates between rNTPs and dNTPs because its nucleotide binding pocket cannot accommodate a 2′-OH on the incoming nucleotide.

Biological Significance

This discrimination prevents the incorporation of ribonucleotides into DNA, which would otherwise destabilize the DNA double helix and increase mutation rates. The structural specificity of DNA polymerase thus ensures genome stability and faithful replication.

Summary Table

Conclusion

DNA polymerase’s ability to discriminate between ribonucleotides and deoxyribonucleotides is primarily due to the structural constraints of its nucleotide binding pocket, which cannot accommodate the 2′-OH group of rNTPs. This ensures that DNA synthesis proceeds with high fidelity, incorporating dNTPs preferentially despite the higher cellular concentration of rNTPs.