Purification and properties of a hyperthermoactive α-amylase from the archaeobacterium Pyrococcus woesei |
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Authors: | Rainhard Koch Andreas Spreinat Karen Lemke Garabed Antranikian |
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Institution: | (1) Arbeitsbereich Biotechnologie I, Technische Mikrobiologie, Technische Universität Hamburg-Harburg, Harburger Schloßstrasse 37, W-2100 Hamburg 90, Federal Republic of Germany;(2) Institut für Mikrobiologie der Georg-August-Universität Göttingen, Grisebachstrasse 8, W-3400 Göttingen, Federal Republic of Germany |
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Abstract: | The cultivation of the hyperthermophilic archaeobacterium Pyrococcus woesei on starch under continuous gassing (80% H2:20% CO2) caused the formation of 250 U/l of an extremely thermoactive and thermostable -amylase. In a complex medium without elemental sulphur under 80% N2 and 20% CO2 atmosphere enzyme production could be elevated up to 1000 U/l. Pyrococcus woesei grew preferentially on poly-and oligosaccharides. The amylolytic enzyme formation was constitutive. Enzyme production was also observed in continuous culture at dilution rates from 0.1 to 0.4 h-1. A 20-fold enrichment of -amylase was achieved after adsorption of the enzyme onto starch and its desorption by preparative gel electrophoresis. The -amylase consisted of a single subunit with a molecular mass of 70 000 and was catalytically active at a temperature range between 40°C and 130°C. Enzymatic activity was detected even after autoclaving at a pressure of 2 bars at 120°C for 5 h. The purified enzyme hydrolyzed exclusively -1,4-glycosidic linkages present in glucose polymers of various sizes. Unlike many -amylases from anaerobes the enzyme from P. woesei was unable to attack short chain oligosaccharides with a chain length between 2 and 6 glucose units. |
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Keywords: | Hyperthermophilic bacteria Thermostability Alpha-amylase Pyrococcus woesei |
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