title: Thiol protection in membrane protein purifications: a study with phage holins.
abstract: The lambda holin, or S105, is a small cytoplasmic membrane protein that controls the timing of host lysis. Using thiol-specific reagents, we determined that the single cysteine residue within S105 was heterogeneously modified during membrane extraction and subsequent immobilized metal ion chromatography. Here we describe the use of a specific and reversible thiol reagent, 2,2'-dithiodipyridine, to generate purified protein with its cysteine residues in the native thiol state. The 2,2'-dithiodipyridine protection protocol was also successfully used for another unrelated holin, S(21)68, and should be generally useful for the purification of membrane proteins.
authors.affiliation: Department of Biochemistry and Biophysics, Texas A&M University, College Station, TX 77843, USA.
authors.forename: Jill S
authors.initials: JS
authors.lastname: Dewey
authors.forename: Douglas K
authors.initials: DK
authors.lastname: Struck
authors.forename: Ry
authors.initials: R
authors.lastname: Young
mesh_terms: D015082:2,2'-Dipyridyl / Q000031:analogs & derivatives*; D000595:Amino Acid Sequence; D001435:Bacteriophages / Q000737:chemistry*; D003545:Cysteine / Q000737:chemistry*; D004220:Disulfides; D008565:Membrane Proteins / Q000737:chemistry / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D014764:Viral Proteins / Q000737:chemistry / Q000302:isolation & purification*
chemical_list: D004220:Disulfides; D008565:Membrane Proteins; D014764:Viral Proteins; C082001:bacteriophage lambda lysis effector protein S105; C001105:2,2'-dipyridyl disulfide; D015082:2,2'-Dipyridyl; D003545:Cysteine