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-d4) 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.
Gu
Sheng
S
BN-2, MS M888, Bioscience Division, Los Alamos National Laboratory, Los Alamos, New Mexico, 87545, USA.
Pan
Songqin
S
Bradbury
E Morton
EM
Chen
Xian
X
D000465:Algorithms; D000595:Amino Acid Sequence; D004268:DNA-Binding Proteins / Q000737:chemistry; D016208:Databases, Factual; D003903:Deuterium; D006801:Humans; D006868:Hydrolysis; D007553:Isotope Labeling; D008239:Lysine / Q000032:analysis*; D008969:Molecular Sequence Data; D018006:Myogenic Regulatory Factors / Q000737:chemistry; D010449:Peptide Mapping; D010455:Peptides / Q000032:analysis*; D011506:Proteins / Q000032:analysis*; D017421:Sequence Analysis; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014157:Transcription Factors; D014357:Trypsin
C503805:ARID5B protein, human; D004268:DNA-Binding Proteins; D018006:Myogenic Regulatory Factors; D010455:Peptides; D011506:Proteins; D014157:Transcription Factors; D003903:Deuterium; D014357:Trypsin; D008239:Lysine