Purification and characterization of aspartate aminotransferase from the halophile archaebacterium Haloferax mediterranei
| Autor: | M C Alvarez-Ossorio, Francisco J. G. Muriana, Angel M. Relimpio |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 1991 |
| Předmět: |
Protein Denaturation
Hot Temperature Chemical Phenomena Macromolecular Substances Biology Biochemistry Transaminase chemistry.chemical_compound Halobacteriaceae Denaturation (biochemistry) Aspartate Aminotransferases Amino Acids Molecular Biology Pyridoxal Chromatography Chemistry Physical Cell Biology Hydrogen-Ion Concentration Phosphate biology.organism_classification Archaea Halophile Haloferax mediterranei Molecular Weight Spectrometry Fluorescence chemistry Spectrophotometry Pyridoxal Phosphate Titration Research Article |
| Popis: | Aspartate aminotransferase from the archaebacterium Haloferax mediterranei was purified and found to be homogeneous. An average Mr of 66,000 was estimated. The native halophilic transaminase exhibited no maximum absorption at 410 nm, which indicates that the apo form is obtained by our purification procedure, and the molar absorption coefficient at 275 nm in 3.5 M-KCl (pH 7.8) was found to be 78.34 mM-1.cm-1. Plots of titration data show that 1 mol of halophilic aspartate aminotransferase binds 2 mol of pyridoxal 5′-phosphate. The halophilic transaminase behaved as a dimer with two similar subunits and had a maximum activity in the pH range 7.6-7.9 and at 65 degrees C in 3.5 M-KCl. By differential scanning calorimetry, the denaturation temperature of the halophilic holo- and apo-transaminase was determined to be 78.5 and 68.0 degrees C respectively at 3.3 M-KCl (pH 7.8). At low salt concentration the halophilic transaminase was inactivated, following first-order kinetics. The Km values for 2-oxoglutarate and L-aspartate, in 3 M-KCl (pH 7.8), were 0.75 mM and 12.6 mM respectively. |
| Databáze: | OpenAIRE |
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