Applied Microbiology and Biotechnology

, Volume 102, Issue 15, pp 6409–6424 | Cite as

Affinity maturation of an antibody for the UV-induced DNA lesions 6,4 pyrimidine-pyrimidones

  • Bingjie Kong
  • Yang Cao
  • Danni Wu
  • Lili An
  • Fanlei Ran
  • Yan Lin
  • Chen Ye
  • Hailin WangEmail author
  • Haiying HangEmail author
Biotechnological products and process engineering


DNA lesions, associated mostly with minor changes in DNA structure, may induce permanent change in heritable coding information. Biochemically, these minor structural changes are difficult to be explored for generating high-affinity antibodies to detect specific DNA lesions in varying sequence contexts. Herein, we established a platform of bacterial display to facilitate antibodies to be matured with high affinity and high specificity against DNA lesions. To achieve this goal, we, for the first time, developed a two-round mutation/screening strategy: (1) using multiple lesion-containing DNA probes for primary maturation and (2) using single lesion-containing DNA probes for second maturation. Specifically, we capitalized on 64M-2 as a parental template to improve affinity for 6-4PP by 710-fold, compared with the model one. In addition, the matured antibody (9c3) is found to be much less dependent on the bases surrounding 6-4PPs than the model one. The mechanistic study from both computational simulation and reverse mutations revealed the critical roles of the two-round mutations in the enhanced binding affinity and independence of surrounding bases. This selection strategy opens a new way to improve affinity and specificity of antibodies for other DNA lesions.


Antibody for DNA lesions Affinity maturation 6,4 Pyrimidine-pyrimidones (6-4PPs) DNA sequence-independent antibody 



We would like to thank Howard B. Lieberman from Columbia University at New York for revising this manuscript, Professor Caixia Guo (Institute of Genomics, Chinese Academy of Sciences) for providing human fibroblasts, Junying Jia and Shuang Sun (Institute of Biophysics, Chinese Academy of Sciences) for their technical assistance in flow cytometry analysis and sorting, and Yuanyuan Chen and Zhenwei Yang (Institute of Biophysics, Chinese Academy of Sciences) for their technical support with Biacore experiments.

Funding information

This work was funded by grants from the Development of Ministry of Science and Technology (No. 2011YQ03013404, No. 2014CB910402) and from the National Natural Science Foundation of China (No. 31500753, No. 31370792, No. 31401130).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

This article does not contain any studies with human participants performed by any of the authors.

Supplementary material

253_2018_8998_MOESM1_ESM.pdf (1.1 mb)
ESM 1 (PDF 1086 kb)


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Bingjie Kong
    • 1
    • 2
  • Yang Cao
    • 3
  • Danni Wu
    • 2
    • 4
  • Lili An
    • 1
  • Fanlei Ran
    • 1
  • Yan Lin
    • 5
  • Chen Ye
    • 1
  • Hailin Wang
    • 2
    • 4
    Email author
  • Haiying Hang
    • 1
    • 2
    Email author
  1. 1.Key Laboratory for Protein and Peptide Pharmaceuticals, National Laboratory of Biomacromolecules, Institute of BiophysicsChinese Academy of SciencesBeijingChina
  2. 2.University of Chinese Academy of SciencesBeijingChina
  3. 3.Center of Growth, Metabolism and Aging, Key Lab of Bio-Resources and Eco-Environment of Ministry of Education, College of Life SciencesSichuan UniversityChengduChina
  4. 4.State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental SciencesChinese Academy of SciencesBeijingChina
  5. 5.College of Bee ScienceFujian Agriculture and Forestry UniversityFuzhouChina

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