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Updated: May 22, 2025

Single-Molecule Fluorescence Visualization of DNA Polymerase Dynamics at G-Quadruplexes
Published on: April 4, 2025
Efficient chemical synthesis of mirror-image DNA polymerase Dpo4 assisted by one-pot multi-segment condensation
Miao Wang1, Tingting Cui2, Shuqing Huang1
1Hefei National Research Center for Interdisciplinary Sciences at the Microscale, University of Science and Technology of China, 230027 Hefei, China. scwei@ustc.edu.cn.
Abstract:
The mirror-image DNA polymerase Dpo4 is a valuable enzymatic tool in biomedical research; however, its chemical synthesis has been hindered by low efficiency. Here, we describe an efficient chemical synthesis of D-Dpo4 using a one-pot multi-segment condensation strategy, which was achieved using Fmoc-masked peptide thioester. By minimizing the need for isolating and handling intermediates through one-pot three- and four-segment ligations, we were able to obtain D-Dpo4 with higher efficiency, achieving an improved overall yield of 9.8%, representing a twofold improvement from previous work. Our work provides a practical and streamlined approach for the synthesis of mirror-image proteins with larger sizes and underscores the utility of one-pot multi-segment condensation in chemical protein synthesis.
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