title: Chemoenzymatic Labeling of Proteins for Imaging in Bacterial Cells.
abstract: Reliable methods to determine the subcellular localization of bacterial proteins are needed for the study of prokaryotic cell biology. We describe here a simple and general technique for imaging of bacterial proteins in situ by fluorescence microscopy. The method uses the eukaryotic enzyme N-myristoyltransferase to modify the N-terminus of the protein of interest with an azido fatty acid. Subsequent strain-promoted azide-alkyne cycloaddition allows conjugation of dyes and imaging of tagged proteins by confocal fluorescence microscopy. We demonstrate the method by labeling the chemotaxis proteins Tar and CheA and the cell division proteins FtsZ and FtsA in Escherichia coli. We observe distinct spatial patterns for each of these proteins in both fixed and live cells. The method should prove broadly useful for protein imaging in bacteria.
authors.affiliation: Division of Chemistry and Chemical Engineering, California Institute of Technology , 1200 East California Boulevard, Pasadena, California 91125, United States.
authors.forename: Samuel H
authors.initials: SH
authors.lastname: Ho
authors.affiliation: Division of Chemistry and Chemical Engineering, California Institute of Technology , 1200 East California Boulevard, Pasadena, California 91125, United States.
authors.forename: David A
authors.initials: DA
authors.lastname: Tirrell
mesh_terms: D000217:Acyltransferases / Q000378:metabolism*; D001426:Bacterial Proteins / Q000032:analysis* / Q000737:chemistry*; D003598:Cytoskeletal Proteins / Q000032:analysis* / Q000737:chemistry*; D004926:Escherichia coli / Q000737:chemistry* / Q000166:cytology*; D029968:Escherichia coli Proteins / Q000032:analysis* / Q000737:chemistry*; D008856:Microscopy, Fluorescence; D015394:Molecular Structure
chemical_list: D001426:Bacterial Proteins; D003598:Cytoskeletal Proteins; D029968:Escherichia coli Proteins; C489310:FtsA protein, E coli; C071525:FtsZ protein, Bacteria; D000217:Acyltransferases; C050480:glycylpeptide N-tetradecanoyltransferase