Large-scale identification of N-terminal peptides in the halophilic archaea Halobacterium salinarum and Natronomonas pharaonis.
Characterization of protein N-terminal peptides supports the quality assessment of data derived from genomic sequences (e.g., the correct assignment of start codons) and hints to in vivo N-terminal modifications such as N-terminal acetylation and removal of the initiator methionine. The current work represents the first large-scale identification of N-terminal peptides from prokaryotes, of the two halophilic euryarchaeota Halobacterium salinarum and Natronomonas pharaonis. Two methods were used that specifically allow the characterization of protein N-terminal peptides: combined fractional diagonal chromatography (COFRADIC) and strong cation exchange chromatography (SCX), both known to enrich for N-terminally blocked peptides. In addition to these specific methods, N-terminal peptide identifications were extracted from our previous genome-wide proteomic data. Combining all data, 606 N-terminal peptides from Hbt. salinarum and 328 from Nmn. pharaonis were reliably identified. These results constitute the largest available dataset holding identified and characterized protein N-termini for prokaryotes (archaea and bacteria). They allowed the validation/improvement of start codon assignments as automatic gene finders tend to misassign start codons for GC-rich genomes. In addition, the dataset allowed unravelling N-terminal protein maturation in archaea, showing that 60% of the proteins undergo methionine cleavage and that-in contrast to current knowledge-Nalpha-acetylation is common in the archaeal domain of life with 13-18% of the proteins being Nalpha-acetylated. The protein sets described in this paper are available by FTP and might be used as reference sets to test the performance of new gene finders.
Aivaliotis
Michalis
M
Department of Membrane Biochemistry, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany.
Gevaert
Kris
K
Falb
Michaela
M
Tebbe
Andreas
A
Konstantinidis
Kosta
K
Bisle
Birgit
B
Klein
Christian
C
Martens
Lennart
L
Staes
An
A
Timmerman
Evy
E
Van Damme
Jozef
J
Siedler
Frank
F
Pfeiffer
Friedhelm
F
Vandekerckhove
Joël
J
Oesterhelt
Dieter
D
D019843:Archaeal Proteins / Q000032:analysis* / Q000235:genetics; D002845:Chromatography / Q000379:methods; D002852:Chromatography, Ion Exchange / Q000379:methods; D019766:Genes, Archaeal; D006216:Halobacteriaceae / Q000737:chemistry* / Q000235:genetics; D019617:Halobacterium salinarum / Q000737:chemistry* / Q000235:genetics; D010455:Peptides / Q000032:analysis; D040901:Proteomics; D020539:Sequence Analysis, Protein
D019843:Archaeal Proteins; D010455:Peptides