title: Determination of deuterium-labeled tryptophan in proteins by means of high-performance liquid chromatography and thermospray mass spectrometry.
abstract: In order to develop direct methods for determining the extent of metabolic incorporation of isotopically labeled amino acids into a protein, the determination of deuterated tryptophan in [2H5]tryptophan-bacteriorhodopsin was investigated. The isotopically modified protein was subjected to alkaline hydrolysis. After phenyl isothiocyanate derivatization of the hydrolysate, the mixture was separated by reversed-phase liquid chromatography. Field desorption mass spectrometry and thermospray mass spectrometry were investigated for their ability to determine the ratio between [2H5]tryptophan and total tryptophan in the collected fractions. In order to check the procedure a set of known tryptophan/[2H5]tryptophan mixtures were passed through the same derivatization, HPLC separation, and lyophilization procedure as used for the biological samples.
authors.affiliation: Department of Organic Chemistry, Leiden University, The Netherlands.
authors.forename: E
authors.initials: E
authors.lastname: Karnaukhova
authors.forename: W M
authors.initials: WM
authors.lastname: Niessen
authors.forename: U R
authors.initials: UR
authors.lastname: Tjaden
authors.forename: J
authors.initials: J
authors.lastname: Raap
authors.forename: J
authors.initials: J
authors.lastname: Lugtenburg
authors.forename: J
authors.initials: J
authors.lastname: van der Greef
mesh_terms: D000596:Amino Acids / Q000032:analysis; D002851:Chromatography, High Pressure Liquid; D003903:Deuterium; D008401:Gas Chromatography-Mass Spectrometry; D006217:Halobacterium / Q000378:metabolism; D017879:Isothiocyanates; D007553:Isotope Labeling; D011506:Proteins / Q000032:analysis*; D013861:Thiocyanates; D014364:Tryptophan / Q000032:analysis*
chemical_list: D000596:Amino Acids; D017879:Isothiocyanates; D011506:Proteins; D013861:Thiocyanates; C005441:phenylisothiocyanate; D014364:Tryptophan; D003903:Deuterium