See & Catch method for studying protein complexes in yeast cells: a technique unifying fluorescence microscopy and mass spectrometry.
We have developed a method for studying proteins and protein complexes in yeast cells based on unification of fluorescence microscopy and mass spectrometry techniques. To apply the method, termed by us as "See & Catch," we first produced a variety of DNA plasmids used as PCR templates for genomic tagging of proteins with a modular fluorescent and affinity tags. The modular tag consists of one of the multiple versions of monomeric fluorescent proteins fused to a variety of small affinity epitopes. Among those modular tags, we found several combinations which were optimal for determining protein subcellular localization and for purifying the tagged proteins and protein complexes for detailed analysis by mass spectrometry. Combining fluorescence microscopy and mass spectrometry into a single method provides a unique possibility to obtain a unified view of the processes regulating dynamic properties of the proteins and protein complexes in living cells.
Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, CA, 94143, USA.
Changhui
C
Deng
Andrew N
AN
Krutchinsky
D002846:Chromatography, Affinity; D013058:Mass Spectrometry; D008856:Microscopy, Fluorescence; D008967:Molecular Biology / Q000379:methods*; D046912:Multiprotein Complexes / Q000737:chemistry; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000235:genetics*
D046912:Multiprotein Complexes; D011993:Recombinant Fusion Proteins