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Published on: January 30, 2019
Determinants of sequence-specificity within human AID and APOBEC3G
Michael A Carpenter1, Erandi Rajagurubandara, Priyanga Wijesinghe
1Department of Chemistry, Wayne State University, Detroit, MI 48202, USA.
DNA Repair
|March 27, 2010
Summary
Human APOBEC3G (A3G) and activation-induced deaminase (AID) are DNA-cytosine deaminases. A key A3G region dictates sequence specificity, while AID sequences in A3G relax this selectivity.
Area of Science:
- Biochemistry
- Molecular Biology
- Genetics
Background:
- Human APOBEC3G (A3G) and activation-induced deaminase (AID) are DNA-cytosine deaminases with distinct sequence preferences.
- A3G targets cytosines in C-runs, while AID targets cytosines in WRC motifs.
- Understanding the structural basis of their sequence selectivity is crucial for elucidating their biological functions.
Purpose of the Study:
- To identify the specific regions within A3G and AID responsible for their DNA sequence selectivity.
- To investigate how swapping these regions affects the catalytic activity and sequence preference of A3G and AID.
- To determine the contribution of different protein segments to the DNA sequence recognition mechanism of APOBEC family enzymes.
Main Methods:
- Structural analysis of the A3G carboxyl-terminal catalytic domain (A3G-CTD) to identify potential DNA-interacting regions.
- Engineering of A3G-CTD and AID hybrids by swapping identified regions (region 1 and region 2).
- Expression of hybrid proteins in Escherichia coli and assessment of their sequence selectivity using genetic and biochemical assays for C-to-T mutations.
Main Results:
- Region 2 (10 amino acids) of A3G-CTD was identified as the principal determinant of its sequence specificity.
- Region 1 of A3G-CTD enhanced the target cytosine preference conferred by region 2.
- Introduction of AID sequences into A3G relaxed its sequence specificity, and neither AID region alone or combined conferred A3G's preference.
Conclusions:
- APOBEC family enzyme sequence selectivity is governed by at least two distinct sequence segments.
- Additional protein regions may be involved in the DNA sequence recognition process.
- The findings provide insights into the structure-function relationships of DNA-cytosine deaminases and their roles in genome evolution and disease.
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