Processive AID-catalysed cytosine deamination on single-stranded DNA simulates somatic hypermutation

Area of Science:

• Immunology
• Molecular Biology
• Genetics

Background:

• Activation-induced cytidine deaminase (AID) is crucial for B cell antibody diversification.
• Antibody class switch recombination and somatic hypermutation (SHM) require AID.
• AID catalyzes cytosine (C) to uracil (U) deamination on single-stranded DNA (ssDNA).

Purpose of the Study:

• To investigate the in vitro catalytic activity and substrate specificity of AID.
• To elucidate the mechanism of AID-mediated DNA deamination.
• To correlate in vitro findings with in vivo characteristics of SHM.

Main Methods:

• In vitro biochemical assays using purified AID protein.
• Analysis of DNA sequences for C to U deamination products.
• Assessment of AID binding affinity to ssDNA and DNA duplexes.

Main Results:

• AID preferentially deaminates C within 5' WRC sequences on ssDNA.
• AID performs multiple deaminations on individual DNA strands, not inter-strand hopping.
• AID shows a 15-fold preference for the non-transcribed DNA strand over the transcribed strand.

Conclusions:

• AID's catalytic activity and substrate preference in vitro explain key features of in vivo SHM.
• AID's strong binding to ssDNA, driven by its positive charge, facilitates multiple deaminations.
• The strand preference of AID deamination is relevant to the transcription-dependence of SHM.