title: The role of peptide deformylase in protein biosynthesis: a proteomic study.
abstract: Recently we investigated the influence of classical and emerging antibiotics on the proteome of Bacillus subtilis including in our studies actinonin, a potent novel inhibitor of peptide deformylase. The protein synthesis pattern under actinonin treatment changed so dramatically that a direct comparison to the control pattern was impossible. Dual channel imaging revealed that actinonin treatment caused the majority of newly synthesised proteins to accumulate in spots different from the ones usually observed, indicating a more acidic isoelectric point. Two strategies were used to investigate the nature of the charge shift. In the first place, protein patterns of a conditional peptide deformylase mutant under nonrepressing and repressing conditions were compared. Secondly, several protein pairs excised from two-dimensional (2-D) gels of the peptide deformylase mutant, exponentially growing untreated wild-type and the actinonin treated wild-type were investigated with matrix-assisted laser desorption/ionization and electrospray ionization (ESI) time of flight mass spectrometry (TOF MS) for the existence of N-terminal formylation. Under nonrepressing conditions the mutant protein pattern resembled that of the wild-type. The loss of peptide deformylase activity under repressing conditions led to the same pI shift observed for actinonin treatment in the wild-type. Quadrupole TOF-MS on 11 protein pairs proved that the remaining N-terminal formyl residue was indeed responsible for the charge shift. Eight of these protein pairs were also present on 2-D gels of exponentially growing B. subtilis, where the more acidic, still formylated protein species represented the smaller parts.
authors.affiliation: Institut für Mikrobiologie, Ernst-Moritz-Arndt-Universität, Greifswald, Germany. julia.bandow@pfizer.com
authors.forename: Julia Elisabeth
authors.initials: JE
authors.lastname: Bandow
authors.forename: Dörte
authors.initials: D
authors.lastname: Becher
authors.forename: Knut
authors.initials: K
authors.lastname: Büttner
authors.forename: Falko
authors.initials: F
authors.lastname: Hochgräfe
authors.forename: Christoph
authors.initials: C
authors.lastname: Freiberg
authors.forename: Heike
authors.initials: H
authors.lastname: Brötz
authors.forename: Michael
authors.initials: M
authors.lastname: Hecker
mesh_terms: D000581:Amidohydrolases / Q000502:physiology*; D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001412:Bacillus subtilis / Q000378:metabolism; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000737:chemistry*; D003600:Cytosol / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D006877:Hydroxamic Acids / Q000494:pharmacology; D007526:Isoelectric Point; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D017434:Protein Structure, Tertiary; D020543:Proteome; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013997:Time Factors
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D006877:Hydroxamic Acids; D010455:Peptides; D020543:Proteome; D000581:Amidohydrolases; C084507:peptide deformylase; C010531:actinonin