Abstract

We propose to investigate certain specific aspects of DNA transposition and excision in bacteriophage Mu. We would like to follow up our surprising observation that transposition of Mu can occur via two pathways and that one of the factors that influence the choice of the pathways could be the proportion of the two proteins A and B that are required for transposition. We will exploit this observation to study specifically one or the other pathway. Besides its involvement in DNA transposition, the A protein also brings about the excision of Mu DNA sequences from their integrated sites. We have cloned the A gene under the strong lacUV5 promoter. Upon overproduction of A, we can detect in vivo, linear excised Mu DNA. We propose to use this observation to test whether excised Mu DNA is an intermediate in transposition. We also plan to reproduce the excision reaction in vitro and thus have an assay to purify the A protein. Eventually we plan to obtain pure transposition proteins in sufficient amounts to study the physico-chemical interactions between the proteins and their substrates.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM033247-03
Application #
3282702
Study Section
Microbial Physiology and Genetics Subcommittee 2 (MBC)
Project Start
1984-04-01
Project End
1988-02-29
Budget Start
1986-04-01
Budget End
1988-02-29
Support Year
3
Fiscal Year
1986
Total Cost
Indirect Cost

  Institution

Name
Scripps Research Institute
Department
Type
DUNS #
City
San Diego
State
CA
Country
United States
Zip Code
92037

  Publications

Jang, Sooin; Harshey, Rasika M (2015) Repair of transposable phage Mu DNA insertions begins only when the E.?coli replisome collides with the transpososome. Mol Microbiol 97:746-58
Harshey, Rasika M (2014) Transposable Phage Mu. Microbiol Spectr 2:
Choi, Wonyoung; Saha, Rudra P; Jang, Sooin et al. (2014) Controlling DNA degradation from a distance: a new role for the Mu transposition enhancer. Mol Microbiol 94:595-608
Choi, Wonyoung; Jang, Sooin; Harshey, Rasika M (2014) Mu transpososome and RecBCD nuclease collaborate in the repair of simple Mu insertions. Proc Natl Acad Sci U S A 111:14112-7
Saha, Rudra P; Lou, Zheng; Meng, Luke et al. (2013) Transposable prophage Mu is organized as a stable chromosomal domain of E. coli. PLoS Genet 9:e1003902
Jang, Sooin; Sandler, Steven J; Harshey, Rasika M (2012) Mu insertions are repaired by the double-strand break repair pathway of Escherichia coli. PLoS Genet 8:e1002642
Lee, Jaemin; Harshey, Rasika M (2012) Loss of FlhE in the flagellar Type III secretion system allows proton influx into Salmonella and Escherichia coli. Mol Microbiol 84:550-65
Harshey, Rasika M (2012) The Mu story: how a maverick phage moved the field forward. Mob DNA 3:21
Lazova, Milena D; Butler, Mitchell T; Shimizu, Thomas S et al. (2012) Salmonella chemoreceptors McpB and McpC mediate a repellent response to L-cystine: a potential mechanism to avoid oxidative conditions. Mol Microbiol 84:697-711
Ge, Jun; Lou, Zheng; Cui, Hong et al. (2011) Analysis of phage Mu DNA transposition by whole-genome Escherichia coli tiling arrays reveals a complex relationship to distribution of target selection protein B, transcription and chromosome architectural elements. J Biosci 36:587-601

Showing the most recent 10 out of 46 publications