Thermostable beta-galactosidase from the archaebacterium Sulfolobus solfataricus. Purification and properties.
A thermophilic and thermostable beta-galactosidase activity was purified to homogeneity from crude extracts of the archaebacterium Sulfolobus solfataricus, by a procedure including ion-exchange and affinity chromatography. The homogeneous enzyme had a specific activity of 116.4 units/mg at 75 degrees C with o-nitrophenyl beta-galactopyranoside as substrate. Molecular mass studies demonstrated that the S. solfataricus beta-galactosidase was a tetramer of 240 +/- 8 kDa composed of similar or identical subunits. Comparison of the amino acid composition of beta-galactosidase from S. solfataricus with that from Escherichia coli revealed a lower cysteine content and a lower Arg/Lys ratio in the thermophilic enzyme. A rabbit serum, raised against the homogeneous enzyme did not cross-react with beta-galactosidase from E. coli. The enzyme, characterized for its reaction requirements and kinetic properties, showed a thermostability and thermophilicity notably greater than those reported for beta-galactosidases from other mesophilic and thermophilic sources.
Pisani
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FM
Istituto di Biochimica delle Proteine ed Enzimologia, Consiglio Nazionale delle Ricerche, Napoli, Italia.
Rella
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Raia
C A
CA
Rozzo
C
C
Nucci
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Gambacorta
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A
De Rosa
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M
Rossi
M
M
D000596:Amino Acids / Q000302:isolation & purification; D001105:Archaea / Q000201:enzymology*; D001419:Bacteria / Q000201:enzymology*; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D003432:Cross-Linking Reagents; D004586:Electrophoresis / Q000379:methods; D004795:Enzyme Stability; D005696:Galactosidases / Q000302:isolation & purification*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007120:Immunochemistry; D007525:Isoelectric Focusing; D007700:Kinetics; D001616:beta-Galactosidase / Q000302:isolation & purification*
D000596:Amino Acids; D003432:Cross-Linking Reagents; D005696:Galactosidases; D001616:beta-Galactosidase