Oligosaccharyltransferase is highly specific for the hydroxy amino acid in Asn-Xaa-Thr/Ser.
Pig liver oligosaccharyltransferase (OST), which is involved in the en bloc transfer of the Dol-PP-linked GlcNAc(2)-Man(9)-Glc(3) precursor on to asparagine residues in the Asn-Xaa-Thr/Ser sequence, is highly stereospecific for the conformation of the 3-carbon atom in the hydroxy amino acid. Moreover, substitution of the hydroxy group by either SH as in cysteine, or NH(2) as in beta,gamma-diamino-butanoic acid as reported previously [Bause, E. et al., Biochem. J. 312 (1995) 979-985], followed by the determination of the pH optimum for enzymatic activity, indicates that neither a negative nor a positive charge in the hydroxy amino acid position is tolerated by the enzyme. Binding of the threonine beta-methyl group by OST is also specific, with serine, L-threo-beta-hydroxynorvaline and L-beta-hydroxynorleucine containing tripeptides all bound much less efficiently than the threonine peptide itself. The data are interpreted in terms of a highly stereospecific hydrophobic binding pocket for the threonine CH(3)-CH(OH) group.
Breuer
W
W
Institut für Physiologische Chemie, Nussallee 11, 53115 Bonn, Germany.
Klein
R A
RA
Hardt
B
B
Bartoschek
A
A
Bause
E
E
D000818:Animals; D001665:Binding Sites; D002384:Catalysis; D006031:Glycosylation; D006602:Hexosyltransferases; D008565:Membrane Proteins; D008968:Molecular Conformation; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*; D013379:Substrate Specificity; D013552:Swine; D014166:Transferases / Q000378:metabolism*
D008565:Membrane Proteins; D010455:Peptides; D014166:Transferases; D006602:Hexosyltransferases; C025620:dolichyl-diphosphooligosaccharide - protein glycotransferase