Probing the proteasome cavity in three steps: bio-orthogonal photo-reactive suicide substrates.
Tri-functional activity-based protein probes that encompass an electrophilic trap, a photo-reactive group and a bio-orthogonal ligation handle are described. With these, and in a three-step chemical proteomics approach, proteasomal catalytic sites are covalently and irreversibly modified, followed by photocrosslinking of these to flanking subunits and Staudinger-Bertozzi ligation for visualization and identification of the resulting conjugates.
Geurink
Paul P
PP
Leiden Institute of Chemistry and Netherlands Proteomics Centre, Gorlaeus Laboratories, Einsteinweg 55, 2333 CC Leiden, The Netherlands.
Florea
Bogdan I
BI
Van der Marel
Gijs A
GA
Kessler
Benedikt M
BM
Overkleeft
Herman S
HS
D000818:Animals; D020134:Catalytic Domain; D045744:Cell Line, Tumor; D051379:Mice; D015335:Molecular Probes / Q000737:chemistry; D055668:Photochemical Processes; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*
D015335:Molecular Probes; D046988:Proteasome Endopeptidase Complex