title: A tris (2-carboxyethyl) phosphine (TCEP) related cleavage on cysteine-containing proteins.
abstract: Introduced in the late 1980s as a reducing reagent, Tris (2-carboxyethyl) phosphine (TCEP) has now become one of the most widely used protein reductants. To date, only a few studies on its side reactions have been published. We report the observation of a side reaction that cleaves protein backbones under mild conditions by fracturing the cysteine residues, thus generating heterogeneous peptides containing different moieties from the fractured cysteine. The peptide products were analyzed by high performance liquid chromatography and tandem mass spectrometry (LC/MS/MS). Peptides with a primary amine and a carboxylic acid as termini were observed, and others were found to contain amidated or formamidated carboxy termini, or formylated or glyoxylic amino termini. Formamidation of the carboxy terminus and the formation of glyoxylic amino terminus were unexpected reactions since both involve breaking of carbon-carbon bonds in cysteine.
authors.affiliation: Department of Biologics Product and Process Development, Bristol-Myers Squibb Technical Operations, Princeton, New Jersey 08543, USA.
authors.forename: Peiran
authors.initials: P
authors.lastname: Liu
authors.forename: Brian W
authors.initials: BW
authors.lastname: O'Mara
authors.forename: Bethanne M
authors.initials: BM
authors.lastname: Warrack
authors.forename: Wei
authors.initials: W
authors.lastname: Wu
authors.forename: Yunping
authors.initials: Y
authors.lastname: Huang
authors.forename: Yihong
authors.initials: Y
authors.lastname: Zhang
authors.forename: Rulin
authors.initials: R
authors.lastname: Zhao
authors.forename: Mei
authors.initials: M
authors.lastname: Lin
authors.forename: Michael S
authors.initials: MS
authors.lastname: Ackerman
authors.forename: Peter K
authors.initials: PK
authors.lastname: Hocknell
authors.forename: Guodong
authors.initials: G
authors.lastname: Chen
authors.forename: Li
authors.initials: L
authors.lastname: Tao
authors.forename: Siegfried
authors.initials: S
authors.lastname: Rieble
authors.forename: Jack
authors.initials: J
authors.lastname: Wang
authors.forename: David B
authors.initials: DB
authors.lastname: Wang-Iverson
authors.forename: Adrienne A
authors.initials: AA
authors.lastname: Tymiak
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Grace
authors.forename: Reb J
authors.initials: RJ
authors.lastname: Russell
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003545:Cysteine / Q000737:chemistry* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism; D010720:Phosphines / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D003545:Cysteine; D010446:Peptide Fragments; D010720:Phosphines; D011506:Proteins; C080938:tris(2-carboxyethyl)phosphine