title: Sequential activation and deactivation of protein function using spectrally differentiated caged phosphoamino acids.
abstract: Photolabile caging groups, including the 1-(2-nitrophenyl)ethyl (NPE) group, have been applied to probe many biological processes, including protein phosphorylation. Although studies with NPE-caged phosphoamino acids have provided valuable information, these investigations have been limited to the use of only one caged species in a single experiment. To expand the scope of these tools, we have developed an approach for sequentially uncaging two different phosphopeptides in one system, enabling interrogation of multiple phosphorylation events. We present the synthesis of [7-(diethylamino)coumarin-4-yl]methyl (DEACM)-caged phosphorylated serine, threonine, and tyrosine building blocks for Fmoc-based solid-phase peptide synthesis to allow convenient incorporation of these residues into peptides and proteins. Exposure of DEACM- and NPE-caged phosphopeptides to 420 nm light selectively releases the DEACM group without affecting the NPE-caged peptide. This then enables a subsequent irradiation event at 365 nm to remove the NPE group and liberate a second phosphopeptide. We demonstrate the versatility of this general sequential uncaging approach by applying it to control Wip1 phosphatase with two wavelengths of light.
authors.affiliation: Department of Chemistry, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, Massachusetts 02139, USA.
authors.forename: Brenda N
authors.initials: BN
authors.lastname: Goguen
authors.forename: Andreas
authors.initials: A
authors.lastname: Aemissegger
authors.forename: Barbara
authors.initials: B
authors.lastname: Imperiali
mesh_terms: D008027:Light; D010455:Peptides / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism; D019606:Phosphoamino Acids / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010766:Phosphorylation; D011506:Proteins / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism
chemical_list: D010455:Peptides; D019606:Phosphoamino Acids; D011506:Proteins; D010749:Phosphoprotein Phosphatases