Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz
Gewählte Publikation:
Hüwel, S; Haalck, L; Conrath, N; Spener, F.
Maltose phosphorylase from Lactobacillus brevis: purification, characterization, and application in a biosensor for ortho-phosphate.
Enzyme Microb Technol. 1997; 21(6):413-420
Doi: 10.1016/S0141-0229(97)00014-8
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- Co-Autor*innen der Med Uni Graz
- Spener Friedrich
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- Abstract:
- With the goal to obtain maltose phosphorylase as a tool to determine ortho-phosphate, the enzyme from Lactobacillus brevis was purified to 98% by an expeditious FPLC-aided procedure which included anion exchange chromatography, gel filtration, and hydroxyapatite chromatography. The native maltose phosphorylase had a molecular mass of 196 kDa and consisted of two 88 kDa subunits. In isoelectric focusing two isoforms with pI values of 4.2 and 4.6 were observed. Maximum enzyme activity was obtained at 36 degrees C and pH 6.5 and was independent of pyridoxal 5'-phosphate. The apparent K(m) values with maltose and phosphate as substrates were 0.9 mmol l-1 and 1.8 mmol l-1, respectively. Maltose phosphorylase could be stored in 10 mM phosphate buffer pH 6.5 at 4 degrees C with a loss of activity of only 7% up to 6 months. The stability of the enzyme at high temperatures was enhanced significantly using additives like phosphate, citrate, and imidazole. The purified maltose phosphorylase was used as key enzyme in a phosphate sensor consisting of maltose phosphorylase and glucose oxidase. A detection limit of 0.1 microM phosphate was observed and the sensor response was linear in the range between 0.5 and 10 microM.
- Find related publications in this database (using NLM MeSH Indexing)
- Biosensing Techniques -
- Calibration -
- Chromatography, Ion Exchange -
- Electrophoresis, Polyacrylamide Gel -
- Enzyme Stability -
- Glucosyltransferases - chemistry Glucosyltransferases - isolation and purification
- Hydrogen-Ion Concentration -
- Kinetics -
- Lactobacillus - enzymology
- Phosphates - analysis
- Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization -
- Temperature -
- Find related publications in this database (Keywords)
- maltose phosphorylase
- purification
- stabilization
- biosensor
- ortho-phosphate determination