Biarsenical-tetracysteine

Spagnuolo CC, Vermeij RJ, Jares-Erijman EA (2006) Improved Photostable FRET-compe-tent biarsenical—tetracysteine probes based on fluorinated fluoresceins. J Am Chem Soc 128 12040-12041... 

Kottegoda S, Aoto PC, Sims CE, Allbritton NL (2008) Biarsenical—tetracysteine motif as a fluorescent tag for detection in capillary electrophoresis. Anal Chem 80 5358-5366... 

Spagnuolo, C. C., Vermeij, R. J. and Jares-Erijman, E. A. (2006). Improved photostable FRET-competent biarsenical-tetracysteine probes based on fluorinated flluoresceins. J. Am. Chem. Soc. 128, 12040-3041. 

The Biarsenical-tetracysteine Protein Tag Chemistry and Biological Applications... 

The biarsenical-tetracysteine method was first introduced more than 7 years ago, and further refinements and development of novel applications are still appearing. Within the last few years, biologists have started to exploit the unique features of this system for probing protein trafficking, turnover, localization, and dynamics. This review aims to describe the conception and development of this protein tag and its applications in the biological sciences. 

Fig. 8.1-1 A comparison of the relative sizes of CFP and the biarsenical-tetracysteine complex. The atoms comprising the chromophores are shown in color with the peptide backbone depicted in green.

The small size of the biarsenical-tetracysteine tag has made it useful in biological studies requiring a genetically encoded fluorescent tag and GFP is not tolerated or has deleterious effects because of its size. FlAsH binding occurs rapidly, does not require any protein secondary structure to generate fluorescence (unlike GFP and its variants that can take minutes to days to become fully fluorescent) and should therefore be a more faithful reporter of the initiation of protein synthesis. The following examples include studies in viruses, bacteria, yeast, and mammalian cells and also indicate the broad applicability of the biarsenical-tetracysteine system. 

The Biarsenical-tetracysteine Protein Tag Chemistry and Biological Applications 443 8.1.4.2 Multicolor Pulse-chase Labeling... 

One noticeable feature of the biarsenical-tetracysteine system is the high stability of the complex when formed, with off-rates up to weeks in vitro [8]. 

The high stability of the biarsenical-tetracysteine complex and the sensitivity of its fluorescence polarization to localized protein dynamics complex has recently been used to probe the structure of the pentameric oligomer of phospholamban [17], a key regulator of contractility in the heart. Tetracysteine sites were formed at three internal sites within the n-helical region involved in oligomerization by mutation of existing amino acids at positions 5, 6,... 

Single-molecule Studies Using Biarsenical-tetracysteines... 

I would like to thank all my coworkers over the years in the Tsien and Ellisman labs who have contributed to the development of the biarsenical-tetracysteine method, particularly Roger Tsien for devising the original concepts and for continual input into their improvement. 

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