Analysis of the phosphoproteome of the multicellular bacterium Streptomyces coelicolor A3(2) by protein/peptide fractionation, phosphopeptide enrichment and high-accuracy mass spectrometry.
The serine (Ser)/threonine (Thr)/tyrosine (Tyr) phosphoproteome of exponentially growing Streptomyces coelicolor A3(2) was analysed using the gel-free approaches of preparative IEF for protein fractionation, followed by strong cation exchange peptide fractionation and phosphopeptide enrichment by TiO(2) metal oxide affinity chromatography. Phosphopeptides were identified using LC-ESI-LTQ-Orbitra MS. Forty-six novel phosphorylation sites were identified on 40 proteins involved in gene regulation or signalling, central metabolism, protein biosynthesis, membrane transport and cell division, as well as several of unknown function. In contrast to other studies, Thr phosphorylation appeared to be preferred, with relative levels of Ser, Thr and Tyr phosphorylation of 34, 52 and 14%, respectively. Genes for most of the 40 phosphorylated proteins reside in the central "housekeeping" region of the linear S. coelicolor chromosome, suggesting that in general Ser, Thr and Tyr phosphorylation play a role in regulating essential aspects of metabolism in streptomycetes. A greater number of regulators and putative regulators were also identified compared with other bacterial phosphoproteome studies, potentially reflecting the complex heterotrophic and developmental life style of S. coelicolor. This study is the first analysis of the phosphoproteome of a member of this morphologically complex and industrially important group of microorganisms.
Parker
Jennifer L
JL
Department of Molecular Microbiology, John Innes Centre, Norwich, UK.
Jones
Alexandra M E
AM
Serazetdinova
Liliya
L
Saalbach
Gerhard
G
Bibb
Mervyn J
MJ
Naldrett
Mike J
MJ
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000032:analysis* / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D010748:Phosphopeptides / Q000032:analysis* / Q000235:genetics; D010750:Phosphoproteins / Q000032:analysis* / Q000235:genetics; D020543:Proteome / Q000032:analysis* / Q000235:genetics; D016415:Sequence Alignment; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D048372:Streptomyces coelicolor / Q000737:chemistry*; D053719:Tandem Mass Spectrometry / Q000379:methods*
D001426:Bacterial Proteins; D010748:Phosphopeptides; D010750:Phosphoproteins; D020543:Proteome