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BY-NC-ND 3.0 license Open Access Published by De Gruyter June 2, 2014

Purification and Properties of Potato Phosphorylase Isozymes

  • Kirumakki N. Shivaram

Abstract

Two multiple forms of α-glucan phosphorylase which migrate about half way in polyacryl-amide-gel electrophoresis (named “slow“ and “fast” isozyme), were isolated by combined chromatography and preparative electrophoresis after freezing the tissue from freshly harvested and from sprouting potato tubers respectively. Depending on the primer used for the synthesis reaction their pH optimum varied between 5.2 and 6.0 and the optimum temperature was 30 and 35 °C. The isoelectric point for the slow isozyme was at pH 5.0±0.1 and for the fast isozyme, pH 5.5±0.1, the molecular weights were 209 000±10 000 and 165 000±5000 and for their subunits 104 000±4000 and 40 000±2000 respectively. Both isozymes were inhibited by Hg2+, Ag+ and p-chloromercuro-benzoate (p-CMB). Fe2+ ions inhibited them partially. Mg2+, Mn2+ and sulfhydryl compounds activated both.

Km values for the slow and fast isozymes with glucose-l-phosphate in presence of soluble starch was 6.7 and 8.0 mM, of amylose 14.3 and 20.0 mм and of glycogen 22.2 and 40.0 mм respectively. The affinity of the primer for the slow and the fast isozymes were as follows: soluble starch 0.5 and 1.0 mм, amylose 2.6 and 3.8 mм, glycogen 6.2 and 7.7 mм respectively. Km values of phosphorolysis with soluble starch was 0.8 and 0.5 mм, with amylose was 3.1 and 1.1 mм, and with glycogen was 6.5 and 1.3 mм respectively. As substrate and primer the soluble starch was superior and the glycogen was inferior. Amylose was in between.

Kinetic parameters suggested the existence of α-glucan phosphorylase isozymes with different specificities: the slow one being more active in the direction of starch synthesis and the fast isozyme degrading faster the polyglucans. These observations suggest that the polyglucan synthesis and degradation in potato tubers may be regulated by the change in the proportion of slow and fast isozymes.

Received: 1976-4-5
Published Online: 2014-6-2
Published in Print: 1976-8-1

© 1946 – 2014: Verlag der Zeitschrift für Naturforschung

This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

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