Charting the proteomes of organisms with unsequenced genomes by MALDI-quadrupole time-of-flight mass spectrometry and BLAST homology searching.
MALDI-quadrupole time-of-flight mass spectrometry was applied to identify proteins from organisms whose genomes are still unknown. The identification was carried out by successively searching a sequence database-first with a peptide mass fingerprint, then with a packet of noninterpreted MS/MS spectra, and finally with peptide sequences obtained by automated interpretation of the MS/MS spectra. A "MS BLAST" homology searching protocol was developed to overcome specific limitations imposed by mass spectrometric data, such as the limited accuracy of de novo sequence predictions. This approach was tested in a small-scale proteomic project involving the identification of 15 bands of gel-separated proteins from the methylotrophic yeast Pichia pastoris, whose genome has not yet been sequenced and which is only distantly related to other fungi.
Shevchenko
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A
Peptide and Protein Group, European Molecular Biology Laboratory, Heidelberg, Germany. shevchenko@EMBL-Heidelberg.de
Sunyaev
S
S
Loboda
A
A
Shevchenko
A
A
Bork
P
P
Ens
W
W
Standing
K G
KG
D000465:Algorithms; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D016208:Databases, Factual; D004285:Dogs; D016681:Genome, Fungal; D007668:Kidney / Q000166:cytology; D008565:Membrane Proteins / Q000737:chemistry; D008969:Molecular Sequence Data; D010449:Peptide Mapping / Q000295:instrumentation / Q000379:methods; D010843:Pichia / Q000235:genetics*; D020543:Proteome / Q000737:chemistry*; D020539:Sequence Analysis, Protein / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014357:Trypsin / Q000378:metabolism
D008565:Membrane Proteins; D020543:Proteome; D014357:Trypsin