{{Short description|Self-labeling protein tag}} thumb|500px|SNAP-tag reaction scheme '''SNAP-tag®''' is a self-labeling protein tag commercially available in various expression vectors. SNAP-tag is a 182 residue polypeptide (19.4 kDa) that can be fused to any protein of interest and further specifically and covalently tagged with a suitable ligand, such as a fluorescent dye. Since its introduction, SNAP-tag has found numerous applications in biochemistry and for the investigation of the function and localisation of proteins and enzymes in living cells.<ref name="pmid21924508">{{cite journal |author1=Crivat G |author2=Taraska JW | title = Imaging proteins inside cells with fluorescent tags | journal = Trends in Biotechnology | volume = 30 | issue = 1 | pages = 8–16 |date=January 2012 | pmid = 21924508 | pmc = 3246539 | doi = 10.1016/j.tibtech.2011.08.002}}</ref>
== Applications == Cell biology utilizes tools that allow manipulation and visualization of proteins in living cells. An important example is the use of fluorescent proteins, such as the green fluorescent protein (GFP) or yellow fluorescent protein (YFP). Molecular biology methods allow these fluorescent proteins to be introduced and expressed in living cells as fusion proteins. However, the photo-physical properties of the fluorescent proteins are generally not suited for single-molecule spectroscopy. Fluorescent proteins have, in comparison to commercially available dyes, a much lower fluorescence quantum yield and are quickly destroyed upon excitation with a focused laser beam (photobleaching).
The SNAP-tag® protein is an engineered version of the ubiquitous mammalian enzyme AGT,<ref name="pmid12725859">{{cite journal |author1=Juillerat A |author2=Gronemeyer T |author3=Keppler A |author4=Gendreizig S |author5=Pick H |author6=Vogel H |author7=Johnsson K | title = Directed evolution of O6-alkylguanine-DNA alkyltransferase for efficient labeling of fusion proteins with small molecules in vivo | journal = Chemistry and Biology | volume = 10 | issue = 4 | pages = 313–317 |date=April 2003 | pmid = 12725859 | doi = 10.1016/S1074-5521(03)00068-1|doi-access=free }}</ref> encoded in humans by the O-6-methylguanine-DNA methyltransferase (MGMT) gene. SNAP-tag was obtained using a directed evolution strategy,<ref>{{Cite journal|last1=Mollwitz|first1=Birgit|last2=Brunk|first2=Elizabeth|last3=Schmitt|first3=Simone|last4=Pojer|first4=Florence|last5=Bannwarth|first5=Michael|last6=Schiltz|first6=Marc|last7=Rothlisberger|first7=Ursula|last8=Johnsson|first8=Kai|date=2012-02-07|title=Directed Evolution of the Suicide Protein O6-Alkylguanine-DNA Alkyltransferase for Increased Reactivity Results in an Alkylated Protein with Exceptional Stability|journal=Biochemistry|volume=51|issue=5|pages=986–994|doi=10.1021/bi2016537|pmid=22280500|issn=0006-2960|url=https://infoscience.epfl.ch/record/175336}}</ref> leading to a hAGT variant that accepts ''O''<sup>6</sup>-benzylguanine derivatives instead of repairing alkylated guanine derivatives in damaged DNA.
An orthogonal tag, called CLIP-tag™, was further engineered from SNAP-tag to accept ''O''<sup>2</sup>-benzylcytosine derivatives as substrates, instead of ''O''<sup>6</sup>-benzylguanine.<ref name="pmid18291317 ">{{cite journal |author1=Gautier A |author2=Juillerat A |author3=Heinis C |author4=Corrêa IR Jr |author5=Kindermann M |author6=Beaufils F |author7=Johnsson K | title = An engineered protein tag for multiprotein labeling in living cells | journal = Chemistry and Biology | volume = 15 | issue = 2 | pages = 128–136 |date=February 2008 | pmid = 18291317 | doi = 10.1016/j.chembiol.2008.01.007|doi-access= |url=http://infoscience.epfl.ch/record/125074 }}</ref> Therefore, Clip-tag- and SNAP-tag-fused proteins can be labeled simultaneously in the same cells. A split-SNAP-tag version suitable for protein complementation assay and protein-protein interaction studies was later developed.<ref name="pmid22910969 ">{{cite journal |author1=Mie M |author2=Naoki T |author3=Uchida K |author4=Kobatake E | title =Development of a split SNAP-tag protein complementation assay for visualization of protein-protein interactions in living cells | journal =Analyst | volume =137 | issue =20 | pages =4760–4765 |date=October 2012 | pmid =22910969 | doi =10.1039/c2an35762c |bibcode=2012Ana...137.4760M |s2cid=6217675 }}</ref>
Apart from fluorescence microscopy, SNAP-tag and CLIP-tag have proven useful in the elucidation of numerous biological processes, including the identification of multiprotein complexes using various approaches such as FRET,<ref name="pmid18488035">{{cite journal |author1=Maurel D |author2=Comps-Agrar L |author3=Brock C |author4=Rives ML |author5=Bourrier E |author6=Ayoub MA |author7=Bazin H |author8=Tinel N |author9=Durroux T |author10=Prézeau L |author11=Trinquet E |author12=Pin JP | title = Cell-surface protein-protein interaction analysis with time-resolved FRET and snap-tag technologies: application to GPCR oligomerization | journal = Nature Methods | volume = 5 | issue = 6 | pages = 561–567 |date=June 2008 | pmid = 18488035 | doi = 10.1038/nmeth.1213| pmc=2642604}}</ref> cross-linking,<ref name="pmid18488035"/> proximity ligation assay,<ref name="pmid22664266">{{cite journal |author1=Gu GJ |author2=Friedman M |author3=Jost C |author4=Johnsson K |author5=Kamali-Moghaddam M |author6=Plückthun A |author7=Landegren U |author8=Söderberg O | title=Protein tag-mediated conjugation of oligonucleotides to recombinant affinity binders for proximity ligation | journal=New Biotechnology | volume=30 | issue=2 | pages=144–152 |date=January 2013 | pmid=22664266 | doi = 10.1016/j.nbt.2012.05.005 | url= https://infoscience.epfl.ch/record/185453}}</ref> as well as the purification of insulin secretory granules of distinct age by doing pulse-chase experiments <ref>{{cite journal |last1=Neukam|first1=Martin|last2=Ganß|first2=Katharina|last3=Vasiljević|first3=Jovana|last4=Broichhagen|first4=Johannes|last5=Johnsson|first5=Kai|last6=Kurth|first6=Thomas|last7=Solimena|first7=Michele|title=Purification of time-resolved insulin granules reveals proteomic and lipidomic changes during granule aging |journal=Cell Reports |date=2024 |volume=43 |issue=3 |article-number=113836 |doi=10.1016/j.celrep.2024.113836 |pmid=38421874 |language=en|biorxiv=10.1101/2020.06.03.103770}}</ref> Other application include the measurement of protein half-lives in vivo,<ref name="pmid23129118">{{cite book |author1=Bodor DL |author2=Rodríguez MG |author3=Moreno N |author4=Jansen LE | title=Analysis of protein turnover by quantitative SNAP-based pulse-chase imaging | journal=Current Protocols in Cell Biology | volume=55 | issue=8.8 | pages=Unit8.8 |date=June 2012 | pmid=23129118 | doi = 10.1002/0471143030.cb0808s55 |isbn=978-0-471-14303-1 |hdl=10400.7/614 }}</ref> and small molecule-protein interactions.<ref name="pmid21499265">{{cite journal |author1=Chidley C |author2=Haruki H |author3=Pedersen MG |author4=Muller E |author5=Johnsson K | title=A yeast-based screen reveals that sulfasalazine inhibits tetrahydrobiopterin biosynthesis | journal=Nature Chemical Biology | volume=7 | issue=6 | pages=375–383 |date=June 2011 | pmid=21499265 | doi = 10.1038/nchembio.557 |url=https://infoscience.epfl.ch/record/166269 }}</ref> SNAP-tag® is a registered trademark of New England Biolabs, Inc. CLIP-tag™ is a trademark of New England Biolabs, Inc.
==See also== * Protein tag * HaloTag * SpyTag * Fluorescent proteins
==References== {{reflist|colwidth=35em}}
== Further reading == {{refbegin|colwidth=35em}} *{{cite journal |author1=Keppler A |author2=Kindermann M |author3=Gendreizig S |author4=Pick H |author5=Vogel H |author6=Johnsson K |title=Labeling of fusion proteins of O6-alkylguanine-DNA alkyltransferase with small molecules in vivo and in vitro |journal=Methods |volume=32 |issue=4 |pages=437–444 |year=2004 |pmid=15003606 |doi=10.1016/j.ymeth.2003.10.007 |url=http://infoscience.epfl.ch/record/81373 }} *{{cite journal |author1=Keppler A |author2=Pick H |author3=Arrivoli C |author4=Vogel H |author5=Johnsson K |title=Labeling of fusion proteins with synthetic fluorophores in live cells |journal=Proc. Natl. Acad. Sci. U.S.A. |volume=101 |issue=27 |pages=9955–9959 |year=2004 |pmid=15226507 |doi=10.1073/pnas.0401923101 |pmc=454197|bibcode=2004PNAS..101.9955K |doi-access=free }} *{{cite journal |author1=Juillerat A |author2=Heinis C |author3=Sielaff I |author4=Barnikow J |author5=Jaccard H |author6=Kunz B |author7=Terskikh A |author8=Johnsson K |title=Engineering substrate specificity of O6-alkylguanine-DNA alkyltransferase for specific protein labeling in living cells |journal=ChemBioChem |volume=6 |issue=7 |pages=1263–1269 |year=2005 |pmid=15934048 |doi=10.1002/cbic.200400431 |s2cid=23926764 |url=http://infoscience.epfl.ch/record/81387 }} {{refend}}
== External links == * Darstellung SNAP-Tag und CLIP-Tag (NEB) * ''Self Labeling Protein Tags.'' In: ''Bioforum.'' Jg. 2005, Nr. 6, S. 50-51.
Category:Biochemistry detection methods