Substrate energetics in non-templated DNA synthesis by terminal deoxynucleotidyl transferase.

TdT uses deoxyribonucleoside diphosphates with reduced rates and efficiencies but similar activation energies to dNTPs, indicating γ-phosphate's role in nucleotide discrimination.

• Why it matters: Understanding substrate energetics in non-templated DNA synthesis is crucial for elucidating enzyme mechanisms and improving enzymatic DNA synthesis technologies.
• What they did: The study examined TdT's activity with dNDPs and dNTPs, measuring rates, efficiencies, substrate bias, and activation energies, revealing that γ-phosphate mainly influences nucleotide discrimination.
• The result: Findings suggest γ-phosphate is not necessary for lowering the energetic barrier of polymerization, providing a mechanistic framework for non-templated DNA synthesis and its technological applications.