Open AccessSubstrate specificity of rat liver cytosolic α‐ d ‐mannosidase Novel degradative pathway for oligomannosidic type glycans
Author(s) -
HAEUW JeanFrançis,
STRECKER Gerard,
WIERUSZESKI JeanMichel,
MONTREUIL Jean,
MICHALSKI JeanClaude
Publication year - 1991
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1991.tb16498.x
Subject(s) - alpha (finance) , cytosol , chemistry , oligosaccharide , glycan , stereochemistry , hydrolysis , biochemistry , substrate (aquarium) , enzyme , biology , glycoprotein , medicine , ecology , construct validity , nursing , patient satisfaction
The substrate specificity of rat liver cytosolic neutral α‐ d ‐mannosidase was investigated by in vitro incubation with a crude cytosolic fraction of oligomannosyl oligosaccharides Man 9 GlcNAc, Man 7 GlcNAc, Man 5 GlcNAc I and II isomers and Man 4 GlcNAc having the following structures: Man 9 GlcNAc, Man(α‐2)Man(α‐3)[Man(α1–2)Man(α1–6)]Man(α1–2)Manα1–2)Man(α1–3)]Man(β‐4)GlcNAc; Man 7 GlcNac, Man(α1–3)[Man(α1–2)Man(α1–6)]Man(α1–6)[Man(α1–2)Man(α1–3]Man(β1–4)GlcNAc; Man 5 GlcNAc I, Man(α1–3)[Man(α1–6)]Man(α1–6)Man(α1–3)]Man(β1–4)GlcNAc; Man 5 GlcNAc II, Man(α1–2)Man(α1–2)Man(α1–3)[Man(α1–6)]Man(β1–4)GlcNAc; Man 4 GlcNAc, Man(α1–2)Man(α1–2)Man(α1–3)Man(β1–4)GlcNAc. The different oligosaccharide isomers resulting from α‐ d ‐mannosidase hydrolysis were analyzed by 1 H‐NMR spectroscopy after HPLC separation. The cytosolic α‐ d ‐mannosidase activity is able to hydrolyse all types of α‐mannosidic linkages found in the glycans of the oligomannosidic type, i.e. α‐1,2, α‐1,3 and α‐1,6. Nevertheless the enzyme is highly active on branched Man 9 GlcNAc or Man 5 GlcNAc I oligosaccharides and rather inactive towards the linear Man 4 GlcNAc oligosaccharide. Structural analysis of the reaction products of the soluble α‐ d ‐mannosidase acting on Man 5 GlcNAc I and Man 9 GlcNAc gives Man 3 GlcNAc, Man(α1–6)[Man(α1–3)]Man(β1–4)GlcNAc, and Man 5 GlcNAc II oligosaccharides, respectively. This Man 5 GlcNAc II, Man(α1–2)Man(α1–2)Man(α1–3)[Man(α1–6)]Man(β1–4)GlcNAc, represents the ‘construction’ Man 5 oligosaccharide chain of the dolichol pathway formed in the cytosolic compartment during the biosynthesis of N ‐glycosylprotein glycans. The cytosolic α‐ d ‐mannosidase is activated by Co 2+ , insensitive to 1‐deoxymannojirimycin but strongly inhibited by swainsonine in the presence of Co 2+ ions. The enzyme shows a highly specific action different from that previously described for the lysosomal α‐ d ‐mannosidases [Michalski, J. C., Haeuw, J. F., Wieruszeski, J. M., Montreuil, J. and Strecker, G. (1990) Eur. J. Biochem. 189 , 369–379]. A possible complementarity between cytoslic and lysosomal α‐ d ‐mannosidase activities in the catabolism of N ‐glycosylprotein is proposed.