title: Selective fluorescent labeling of S-nitrosothiols (S-FLOS): a novel method for studying S-nitrosation.
abstract: Protein S-nitrosation is a reversible post-translation modification critical for redox-sensitive cell signaling that is typically studied using the Biotin Switch method. This method and subsequent modifications usually require avidin binding or Western blot analysis to detect biotin labeled proteins. We describe here a modification of the Biotin Switch assay that eliminates the need for Western blot or avidin enrichment protocols and allows direct comparison of the S-nitrosation state proteins from two different samples in the same gel lane or on the same 2D gel. This S-FLOS method offers detection, identification and quantification of S-nitrosated proteins, with the potential for site-specific identification of nitrosation events.
authors.affiliation: The Johns Hopkins School of Medicine, Anesthesiology and CCM, 720 Rutland Avenue, Traylor 621, Baltimore, MD 21205, USA.
authors.forename: Lakshmi
authors.initials: L
authors.lastname: Santhanam
authors.forename: Marjan
authors.initials: M
authors.lastname: Gucek
authors.forename: Tashalee R
authors.initials: TR
authors.lastname: Brown
authors.forename: Malini
authors.initials: M
authors.lastname: Mansharamani
authors.forename: Sungwoo
authors.initials: S
authors.lastname: Ryoo
authors.forename: Christopher A
authors.initials: CA
authors.lastname: Lemmon
authors.forename: Lewis
authors.initials: L
authors.lastname: Romer
authors.forename: Artin A
authors.initials: AA
authors.lastname: Shoukas
authors.forename: Dan E
authors.initials: DE
authors.lastname: Berkowitz
authors.forename: Robert N
authors.initials: RN
authors.lastname: Cole
mesh_terms: D000818:Animals; D001710:Biotin; D001923:Brain Chemistry; D002232:Carbocyanines; D030562:Databases, Protein; D005453:Fluorescence; D008301:Maleimides; D051379:Mice; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000378:metabolism; D015538:Nitrosation; D010084:Oxidation-Reduction; D011499:Protein Processing, Post-Translational; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D040901:Proteomics; D026403:S-Nitrosothiols / Q000032:analysis; D016622:Silver Staining; D053719:Tandem Mass Spectrometry
chemical_list: D002232:Carbocyanines; D008301:Maleimides; D011506:Proteins; D026403:S-Nitrosothiols; C089187:cyanine dye 3; C085321:cyanine dye 5; C043592:maleimide; D009569:Nitric Oxide; D001710:Biotin; D019001:Nitric Oxide Synthase