HiRIEF LC-MS enables deep proteome coverage and unbiased proteogenomics.
We present a liquid chromatography-mass spectrometry (LC-MS)-based method permitting unbiased (gene prediction-independent) genome-wide discovery of protein-coding loci in higher eukaryotes. Using high-resolution isoelectric focusing (HiRIEF) at the peptide level in the 3.7-5.0 pH range and accurate peptide isoelectric point (pI) prediction, we probed the six-reading-frame translation of the human and mouse genomes and identified 98 and 52 previously undiscovered protein-coding loci, respectively. The method also enabled deep proteome coverage, identifying 13,078 human and 10,637 mouse proteins.
1] Department of Oncology-Pathology, Science for Life Laboratory, Karolinska Institutet, Stockholm, Sweden. [2].
Rui M M
RM
Branca
1] Department of Oncology-Pathology, Science for Life Laboratory, Karolinska Institutet, Stockholm, Sweden. [2].
Lukas M
LM
Orre
1] Department of Oncology-Pathology, Science for Life Laboratory, Karolinska Institutet, Stockholm, Sweden. [2].
Henrik J
HJ
Johansson
Department of Biochemistry and Biophysics, Science for Life Laboratory, Stockholm University, Stockholm, Sweden.
Viktor
V
Granholm
Department of Biochemistry and Biophysics, The Arrhenius Laboratories for Natural Sciences, Science for Life Laboratory, Stockholm University, Stockholm, Sweden.
Mikael
M
Huss
Department of Oncology-Pathology, Science for Life Laboratory, Karolinska Institutet, Stockholm, Sweden.
Åsa
Å
Pérez-Bercoff
Department of Oncology-Pathology, Science for Life Laboratory, Karolinska Institutet, Stockholm, Sweden.
Jenny
J
Forshed
1] School of Biotechnology, Science for Life Laboratory, KTH Royal Institute of Technology, Stockholm, Sweden. [2] Swedish e-Science Resource Center, KTH Royal Institute of Technology, Stockholm, Sweden.
Lukas
0000000156899797
L
Käll
Department of Oncology-Pathology, Science for Life Laboratory, Karolinska Institutet, Stockholm, Sweden.
Janne
J
Lehtiö
D000818:Animals; D017360:Arabidopsis / Q000235:genetics; D002853:Chromatography, Liquid / Q000379:methods*; D019295:Computational Biology / Q000379:methods; D005091:Exons; D023281:Genomics / Q000379:methods*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing / Q000379:methods; D013058:Mass Spectrometry / Q000379:methods*; D051379:Mice; D015233:Models, Statistical; D016366:Open Reading Frames; D010455:Peptides / Q000737:chemistry; D014176:Protein Biosynthesis; D011506:Proteins / Q000737:chemistry; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods*
D010455:Peptides; D011506:Proteins; D020543:Proteome