Glycosylation of the phosphate binding protein, PstS, in Streptomyces coelicolor by a pathway that resembles protein O-mannosylation in eukaryotes.
Previously mutations in a putative protein O-mannosyltransferase (SCO3154, Pmt) and a polyprenol phosphate mannose synthase (SCO1423, Ppm1) were found to cause resistance to phage, phiC31, in the antibiotic producing bacteria Streptomyces coelicolor A3(2). It was proposed that these two enzymes were part of a protein O-glycosylation pathway that was necessary for synthesis of the phage receptor. Here we provide the evidence that Pmt and Ppm1 are indeed both required for protein O-glycosylation. The phosphate binding protein PstS was found to be glycosylated with a trihexose in the S. coelicolor parent strain, J1929, but not in the pmt(-) derivative, DT1025. Ppm1 was necessary for the transfer of mannose to endogenous polyprenol phosphate in membrane preparations of S. coelicolor. A mutation in ppm1 that conferred an E218V substitution in Ppm1 abolished mannose transfer and glycosylation of PstS. Mass spectrometry analysis of extracted lipids showed the presence of a glycosylated polyprenol phosphate (PP) containing nine repeated isoprenyl units (C(45)-PP). S. coelicolor membranes were also able to catalyse the transfer of mannose to peptides derived from PstS, indicating that these could be targets for Pmt in vivo.
Wehmeier
S
S
School of Medical Sciences, Institute of Medical Sciences, University of Aberdeen, Aberdeen AB25 2ZD, UK.
Varghese
A S
AS
Gurcha
S S
SS
Tissot
B
B
Panico
M
M
Hitchen
P
P
Morris
H R
HR
Besra
G S
GS
Dell
A
A
Smith
M C M
MC
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D006031:Glycosylation; D008358:Mannose / Q000378:metabolism; D008969:Molecular Sequence Data; D033903:Periplasmic Proteins / Q000235:genetics / Q000378:metabolism*; D028044:Phosphate-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D020539:Sequence Analysis, Protein; D048372:Streptomyces coelicolor / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; D033903:Periplasmic Proteins; D028044:Phosphate-Binding Proteins; D008358:Mannose