Use of Deuterium-Labeled Lysine for Efficient Protein Identification and Peptide de Novo Sequencing

Here, we describe a method for protein identification and de novo peptide sequencing. Through in vivo cell culturing, the deuterium-labeled lysine residue (Lys-d 4) introduces a 4-Da mass tag at the carboxyl terminus of proteolytic peptides when cleaved by certain proteases. The 4-Da mass difference...

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Bibliographic Details
Published in:Analytical chemistry (Washington) 2002-11, Vol.74 (22), p.5774-5785
Main Authors: Gu, Sheng, Pan, Songqin, Bradbury, E. Morton, Chen, Xian
Format: Article
Language:English
Subjects:
Algorithms
Amino Acid Sequence
Analytical, structural and metabolic biochemistry
Biological and medical sciences
Chemistry
Databases, Factual
Deuterium
DNA-Binding Proteins - chemistry
Fundamental and applied biological sciences. Psychology
General aspects, investigation methods
Humans
Hydrolysis
Isotope Labeling
Lysine - analysis
Molecular Sequence Data
Myogenic Regulatory Factors - chemistry
Peptide Mapping
Peptides
Peptides - analysis
Proteins
Proteins - analysis
Scientific imaging
Sequence Analysis
Spectrometry, Mass, Electrospray Ionization
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Transcription Factors
Trypsin
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Summary:Here, we describe a method for protein identification and de novo peptide sequencing. Through in vivo cell culturing, the deuterium-labeled lysine residue (Lys-d 4) introduces a 4-Da mass tag at the carboxyl terminus of proteolytic peptides when cleaved by certain proteases. The 4-Da mass difference between the unlabeled and the deuterated lysine assigns a mass signature to all lysine-containing peptides in any pool of proteolytic peptides for protein identification directly through peptide mass mapping. Furthermore, it was used to distinguish between N- and C-terminal fragments for accurate assignments of daughter ions in tandem MS/MS spectra for sequence assignment. This technique simplifies the labeling scheme and the interpretation of the MS/MS spectra by assigning different series of fragment ions correctly and easily and is very useful in de novo peptide sequencing. We have also successfully implemented this approach to the analysis of protein mixtures derived from the human proteome.
ISSN:0003-2700
1520-6882
DOI:10.1021/ac0204350