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Tracking intracellular protein movements using photoswitchable fluorescent proteins PS-CFP2 and Dendra2.
Nat Protoc 2007; 2(8):2024-32NP

Abstract

A number of photoactivatable GFP-like fluorescent proteins (PAFPs) have been reported whose fluorescence can be switched on or whose fluorescent state can be modified by relatively intense irradiation at a specific wavelength. The use of these proteins gives unique opportunities to photolabel and track fusion proteins in a living cell. Here, we provide a protocol for the primary visualization, photoactivation and tracking of two monomeric PAFPs recently developed in our lab. Both these proteins, PS-CFP2 and Dendra2, are fluorescent and can be visualized before photoactivation. Upon photoactivation, their excitation and emission spectra undergo a dramatic red shift. The brightness of their initial and photoconverted states, along with the high dynamic ranges of both proteins, make them an attractive tool for protein photolabeling. Excluding genetic constructs cloning, cell culturing and transfection, the whole protocol may take anywhere from 10 min to several hours, depending on motility of the protein being studied.

Authors+Show Affiliations

Laboratory of Molecular Technologies for Biology and Medicine, Institute of Bioorganic Chemistry, Russian Academy of Sciences, Miklukho-Maklaya 16/10, Moscow 117997, Russia.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

17703215

Citation

Chudakov, Dmitriy M., et al. "Tracking Intracellular Protein Movements Using Photoswitchable Fluorescent Proteins PS-CFP2 and Dendra2." Nature Protocols, vol. 2, no. 8, 2007, pp. 2024-32.
Chudakov DM, Lukyanov S, Lukyanov KA. Tracking intracellular protein movements using photoswitchable fluorescent proteins PS-CFP2 and Dendra2. Nat Protoc. 2007;2(8):2024-32.
Chudakov, D. M., Lukyanov, S., & Lukyanov, K. A. (2007). Tracking intracellular protein movements using photoswitchable fluorescent proteins PS-CFP2 and Dendra2. Nature Protocols, 2(8), pp. 2024-32.
Chudakov DM, Lukyanov S, Lukyanov KA. Tracking Intracellular Protein Movements Using Photoswitchable Fluorescent Proteins PS-CFP2 and Dendra2. Nat Protoc. 2007;2(8):2024-32. PubMed PMID: 17703215.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Tracking intracellular protein movements using photoswitchable fluorescent proteins PS-CFP2 and Dendra2. AU - Chudakov,Dmitriy M, AU - Lukyanov,Sergey, AU - Lukyanov,Konstantin A, PY - 2007/8/19/pubmed PY - 2007/12/7/medline PY - 2007/8/19/entrez SP - 2024 EP - 32 JF - Nature protocols JO - Nat Protoc VL - 2 IS - 8 N2 - A number of photoactivatable GFP-like fluorescent proteins (PAFPs) have been reported whose fluorescence can be switched on or whose fluorescent state can be modified by relatively intense irradiation at a specific wavelength. The use of these proteins gives unique opportunities to photolabel and track fusion proteins in a living cell. Here, we provide a protocol for the primary visualization, photoactivation and tracking of two monomeric PAFPs recently developed in our lab. Both these proteins, PS-CFP2 and Dendra2, are fluorescent and can be visualized before photoactivation. Upon photoactivation, their excitation and emission spectra undergo a dramatic red shift. The brightness of their initial and photoconverted states, along with the high dynamic ranges of both proteins, make them an attractive tool for protein photolabeling. Excluding genetic constructs cloning, cell culturing and transfection, the whole protocol may take anywhere from 10 min to several hours, depending on motility of the protein being studied. SN - 1750-2799 UR - https://www.unboundmedicine.com/medline/citation/17703215/Tracking_intracellular_protein_movements_using_photoswitchable_fluorescent_proteins_PS_CFP2_and_Dendra2_ L2 - http://dx.doi.org/10.1038/nprot.2007.291 DB - PRIME DP - Unbound Medicine ER -