Escherichia coli beta-galactosidase unexpectedly cleaves the hexasaccharide Gal beta 1-4GlcNAc beta 1-3(Gal beta 1-4GlcNAc beta 1-6)Gal beta 1-4GlcNAc without branch specificity.
The branch specificity of Escherichia coli beta-galactosidase (EC 3.2.1.23) was studied by analyzing the cleavage of the branched hexasaccharide Gal beta 1-4GlcNAc beta 1-3(Gal beta 1-4GlcNAc beta 1-6)[14C(U)]Gal beta 1-4GlcNAc (1). This hexasaccharide was cleaved to pentasaccharides Gal beta 1-4GlcNAc beta 1-3(GlcNAc beta 1-6) [14C(U)]Gal beta 1-4GlcNAc (3) and GlcNAc beta 1-3(Gal-beta 1-4GlcNAc beta 1-6) [14C(U)]Gal beta 1-4GlcNAc (4) without any appreciable branch specificity. Even the further conversions of the pentasaccharides 3 and 4 into the tetrasaccharide GlcNAc beta 1-3(GlcNAc beta 1-6)[14C(U)]Gal beta 1-4GlcNAc seemed to proceed at similar rates, without any appreciable branch specificity. In marked contrast to the hexasaccharide 1, the pentasaccharide Gal beta 1-4GlcNAc beta 1-3(Gal beta 1-4GlcNAc beta 1-6)[14C(U)]Gal (2), missing the reducing end GlcNAc, is known to be cleaved selectively at the 6-branch; this finding was confirmed in the present study. The different behaviour of hexasaccharide 1 and pentasaccharide 2 reflects differences in the reactivity of their 6-branches; the preferred conformations of these closely related molecules may be quite different.
Renkonen
O
O
Institute of Biotechnology, University of Helsinki, Finland.
Helin
J
J
Vainio
A
A
Niemelä
R
R
Penttilä
L
L
Hilden
P
P
D000117:Acetylglucosamine / Q000031:analogs & derivatives*; D002240:Carbohydrate Sequence; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism*; D013379:Substrate Specificity; D001616:beta-Galactosidase / Q000378:metabolism*
D009844:Oligosaccharides; D011134:Polysaccharides; C066360:galactosyl-beta 1-4-N-acetylglucosaminyl-beta 1-3-(galactosyl-beta 1-4-N-acetylglucosaminyl-beta 1-6)galactosyl-beta 1-4-N-acetylglucosaminylpyranoside; D001616:beta-Galactosidase; D000117:Acetylglucosamine