A simple intact protein analysis by MALDI-MS for characterization of ribosomal proteins of two genome-sequenced lactic acid bacteria and verification of their amino acid sequences.
Rapid identification of bacteria by a bioinformatics-based approach, which processes the mass spectra observed by matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS), relies on the calculated masses of ribosomal subunit proteins as biomarkers predicted from amino acid sequences found in protein sequence databases. To verify the actual state of the registered sequence information, a simple intact protein analysis by MALDI-MS using cell lysates as samples was applied to the characterization of ribosomal proteins from genome-sequenced Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus strains. This method avoided the risk of loss of some subunit proteins and the formation of disulfide bonds during the purification of ribosomal proteins. By comparing this with the MALDI mass spectra of different strains and carrying out manual inspection of sequence information, a total of five errors in N-terminal amino acid sequences were identified. After sequence correction, approximately 40 out of 53 subunit proteins could be assigned, considering N-terminal methionine loss only as a post-translational modification. These show promise for use as practical biomarkers for the rapid identification of S. thermophilus and L. bulgaricus. After verification of these amino acid sequences, mass differences relative to those of genome-sequenced strains have the potential for distinguishing bacteria at the strain level.
Teramoto
Kanae
K
Research Institute for Environmental Management Technology, National Institute of Advanced Industrial Science and Technology, Tsukuba, Ibaraki 305-8569, Japan.
Sato
Hiroaki
H
Sun
Liwei
L
Torimura
Masaki
M
Tao
Hiroaki
H
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000032:analysis* / Q000235:genetics; D016680:Genome, Bacterial; D007778:Lactobacillus / Q000737:chemistry*; D008969:Molecular Sequence Data; D009154:Mutation; D012269:Ribosomal Proteins / Q000032:analysis* / Q000235:genetics; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D048249:Streptococcus thermophilus / Q000737:chemistry* / Q000235:genetics
D001426:Bacterial Proteins; D012269:Ribosomal Proteins