Neocarratetraose 4‐O‐Monosulphate β‐Hydrolase from Pseudomonas carrageenovora

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Abstract

Neocarratetraose 4‐O‐monosulphate β‐hydrolase has been enriched 73‐fold from the cell‐free extract of Pseudomonas carrageenovora. This enzyme catalyses the hydrolysis of the β‐glycosidic linkage of neocarratetraose 4‐O‐monosulphate [3,6‐anhydro‐α‐d‐galactopyranosyl‐(1–>3)‐1‐d‐galactopyranosyl‐(1–>4)‐3,6‐anhydro‐α‐d‐galactopyranosyl‐(1–>3)‐ d‐galactose 4‐O‐sulphate]. The products of hydrolysis are respectively neocarrabiose [3,6‐anhydro‐α‐d‐galactopyranosyl‐(1–>3)‐ d‐galactose] and neocarrabiose 4‐O‐sulphate [3,6‐anhydro‐α‐d‐galactopyranosyl‐(1–>3)‐ d‐galactose 4‐O‐sulphate]. During purification the χ‐carrageenase and 4‐sulphatase activities present in the bacterial extract were resolved from the title enzyme. From dodecylsalphate/polyacrylamide gel electrophoresis the enzyme has a molecular weight of 94000. Calcium stimulated the activity, Km 0.50 mmol dm−3. Optimal activity was found at pH 7.0 and the Km determined for the nominal substrate was 68 μmol dm−3. [14C]Neocarratetraose 4‐O‐monosulphate was prepared from [14C]carrageenan (Chondrus crispus) and utilized in a radiochemical assay for neocarratetraose 4‐O‐monosulphate β‐hydrolase. Synthesis of [1‐3H]neocarratetraitol 4‐O‐[35S]monosulphate yielded another substrate. This alditol was hydrolysed to yield a single radioactive species, thus supporting the position of sulphation of neocarratetraose 4‐O‐monosulphate deduced from 13C NMR studies. Neocarratetraose 4‐O‐disulphate [3,6‐anhydro‐α‐d‐galactopyranosyl‐(1–>3)‐4‐O‐sulphato‐β‐d‐galactopyranosyl‐(1–>4)‐3,6‐anhydro‐α‐d‐galactopyranosyl‐(1–>3)‐d‐galactose 4‐O‐sulphate] was not hydrolysed by the purified glycosidase. Tentative evidence for transglycosylation was obtained with both monosulphated substrates. A pathway is described for the sequential degradation of neocarratetraose 4‐O‐disulphate to neocarrabiose. Copyright © 1981, Wiley Blackwell. All rights reserved

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McLEAN, M. W., & WILLIAMSON, F. B. (1981). Neocarratetraose 4‐O‐Monosulphate β‐Hydrolase from Pseudomonas carrageenovora. European Journal of Biochemistry, 113(3), 447–456. https://doi.org/10.1111/j.1432-1033.1981.tb05084.x

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