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Tn5/IS50 target recognition
I Y Goryshin1, J A Miller, Y V Kil
1Department of Biochemistry, University of Wisconsin-Madison, 420 Henry Mall, Madison, WI 53706, USA.
Summary
Transposase protein alone determines Tn5/IS50 transposition target site selection. Analysis reveals a consensus target sequence and suggests multiple transposase interactions influence insertion site clustering and periodicity.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- Transposable elements like Tn5/IS50 play crucial roles in genome evolution.
- Understanding the mechanism of transposition, particularly target site selection, is fundamental to molecular biology.
Purpose of the Study:
- To elucidate the factors governing Tn5/IS50 transposition target site selection.
- To determine if transposase alone mediates target recognition.
- To identify the sequence determinants and higher-order organizational principles of Tn5/IS50 insertion.
Main Methods:
- Analysis of extensive Tn5 and IS50 insertion collections in small DNA regions.
- Comparison of in vitro and in vivo transposition data.
- Deduction of target consensus sequences from insertion sites.
- Design and testing of synthetic target sequences.
Main Results:
- Transposase protein alone dictates target site selection, confirmed by in vitro experiments.
- A consensus Tn5/IS50 target sequence (A-GNTYWRANC-T) was identified.
- Insertion sites exhibit clustering and 5-bp periodicity, suggesting cooperative binding of multiple transposase protomers.
Conclusions:
- Target selection is an intrinsic property of the Tn5/IS50 transposase.
- Cooperative binding of multiple transposase protomers influences insertion site preference and organization.
- The model of Tn5/IS50 target selection is supported by synthetic target sequence analysis.