Chemical tools for protein imaging in live bacterial cells

Abstract

Reliable methods to det. the subcellular localization of bacterial proteins are needed for the study of prokaryotic cell biol. We report a simple and general technique for imaging 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 cycloaddn. allows conjugation of dyes and imaging of tagged proteins by confocal fluorescence microscopy. This method is demonstrated through labeling of the chemotaxis proteins Tar and CheA and the cell division proteins FtsZ and FtsA in Escherichia coli. Distinct spatial patterns for each of these proteins are obsd. in both fixed and live cells. Furthermore, we describe a photoswitchable fluorescent reporter to effect super-resoln. imaging of bacterial chemotaxis proteins. The methods described herein should prove broadly useful for protein imaging in bacteria.