Purification of a membrane-derived proteinase capable of activating a galactosyltransferase involved in volume regulation |
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Authors: | Dorothea Köhle Heinrich Kauss |
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Institution: | Universität Kaiserslautern, Fachbereich Biologie, Postfach 3049, D-6750 Kaiserslautern, F.R.G. |
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Abstract: | UDPgalactose:sn-glycerol-3-phosphate α-D-galactosyltransferase (IFP-synthase, EC 2.4.1.96) shows low activity in extracts prepared from standard volume cells of Poterioochromonals malhamensis under certain conditions. This inactive enzyme has been partially purified by chromatography on DEAE-cellulose, Sephadex G-150 and α-lactalbumin-agarose. It can be activated by an auxiliary enzyme which can be eluted from membranes and which has been purified to homogeneity by chromatography on DEAE-Sephacel and immobilized hemoglobin and fetuin. The activating enzyme is inhibited by chymostatin, antipain and diisopropylfluorophosphate and does not require divalent ions. It consists of a single peptide chain of molecular weight 46 000, can split certain proteins and appears to be a serine proteinase operating around a pH of 6.0. The activating proteinase is irreversibly generated in the crude homogenates on addition of Ca2+ and also shows increased activity shortly after cell shrinkage. This might indicate that it represents one of the possibilities to render the galactosyltransferase active as a result of the physiological stimulus. |
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Keywords: | Proteinase Galactosyltransferase Volume regulation (P malhamensis) IFP-synthase UDPgalactose Hepes 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid Mes 4-morpholinoethanesulfonic acid |
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