The nature of the carbohydrate-peptide linkage region in glycoproteins from the cellulosomes of Clostridium thermocellum and Bacteroides cellulosolvens.
The cellulase complexes of two cellulolytic bacteria, Clostridium thermocellum and Bacteroides cellulosolvens, were subjected to extensive Pronase digestion. Glycopeptide fractions were isolated by gel permeation and fast protein liquid chromatography and analyzed by monosaccharide analysis, amino acid analysis, methylation analysis, and 1H NMR spectroscopy. Alkaline borohydride-induced deglycosylation/amino acid conversion and periodate oxidation studies on the glycopeptide fraction of the C. thermocellum cellulosome demonstrated that the earlier established collection of carbohydrate moieties with 3-O-Me-D-GlcpNAc-alpha (1-->2)-[D-Galp-alpha (1-->3)]-D-Galf-alpha (1-->2)-D-Gal (where 3-O-Me-D-GlcpNAc is 3-O-methyl-N-acetylglucopyranosamine, Galp is galactopyranose, and Galf is galactofuranose) as the major component, is O-linked to threonine via galactopyranose. Using the same approach for the glycopeptide fraction of the cellulase complex of B. cellulosolvens, it was found that the reported collection of carbohydrate moieties with D-Galf-alpha (1-->3)-D-GlcpNAc-alpha (1-->2)-D-Galf-alpha (1-->2)-[D-Galf-beta (1-->3)]-D-Gal as the major component, is O-linked mainly to threonine and partly to serine via galactopyranose. In both species, the hydroxyamino-acid-bound galactopyranose residue has probably an alpha-configuration. The carbohydrate chains appear as clusters located in highly Thr/Pro-rich peptide regions of the glycoproteins. The results are consistent with the notion that the glycosylation sites are localized in linker sequences which connect the various binding domains of the noncatalytic S1 subunit of the cellulosome.
Gerwig
G J
GJ
Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Kamerling
J P
JP
Vliegenthart
J F
JF
Morag
E
E
Lamed
R
R
Bayer
E A
EA
D001439:Bacteroides / Q000737:chemistry*; D002240:Carbohydrate Sequence; D002241:Carbohydrates / Q000737:chemistry*; D002480:Cellulase / Q000737:chemistry*; D003013:Clostridium / Q000737:chemistry*; D006023:Glycoproteins / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry
D002241:Carbohydrates; D006023:Glycoproteins; D010455:Peptides; D002480:Cellulase