A high-throughput assay for DNA deaminases

Methods Mol Biol. 2011:718:171-84. doi: 10.1007/978-1-61779-018-8_11.

Abstract

Most members of the AID/APOBEC family of polynucleotide deaminases can catalyse the deamination of cytosine to uracil in DNA. They thereby function as active DNA mutators. Here, we describe how bacterial papillation assays can be adapted to monitor the mutator activity of AID/APOBEC proteins and show how such papillation assays can be used as a high-throughput screen to identify AID variants with increased specific activity. It should also be possible to use papillation assays for the identification of novel DNA deaminases.

MeSH terms

  • APOBEC-1 Deaminase
  • Cytidine Deaminase / genetics*
  • Cytidine Deaminase / metabolism
  • Cytosine / metabolism
  • DNA / genetics*
  • DNA / metabolism
  • Escherichia coli / genetics*
  • Escherichia coli / isolation & purification
  • Escherichia coli / metabolism
  • Escherichia coli Proteins / genetics*
  • Escherichia coli Proteins / metabolism
  • High-Throughput Screening Assays / methods*
  • Mutation*

Substances

  • Escherichia coli Proteins
  • Cytosine
  • DNA
  • AICDA (activation-induced cytidine deaminase)
  • APOBEC-1 Deaminase
  • Cytidine Deaminase