Purification and characterization of the alcohol dehydrogenase from a novel strain of Bacillus stearothermophilus growing at 70 degrees C.
The biocatalysts isolated from thermophilic microorganisms are the object of ever-growing scientific interest for (i) the comprehension of the molecular basis of their thermal tolerance, and (ii) their use in different bio-industrial fields. Here we report the purification and characterization of an alcohol dehydrogenase (designated ADH-hT) from the novel strain LLD-R of Bacillus stearothermophilus which grows at 70 degrees C. ADH-hT was obtained in pure form by anion exchange chromatography and two affinity chromatographies, with a final yield of about 30%. ADH-hT was found to be a tetramer of 37 kDa-subunits, and to have a pI of 4.9. ADH-hT displayed a broad substrate specificity; its activity was highest for aldehydes, and decreased progressively for alcohols and ketones. ADH-hT was endowed with catalytic activity and resistance in the presence of several denaturing agents (organic solvents, detergents, chaotropic agents). ADH-hT shared with ADH 1503 (the alcohol dehydrogenase from B. stearothermophilus strain NCA 1503 which grows at 55 degrees C) the optimal temperature of 65 degrees C, but it was more resistant than ADH 1503 towards heating. In conclusion, due to its stability and broad substrate specificity ADH-hT could be utilized in bio-industrial processes. Furthermore, we believe that ADH-hT could represent a good model system for studying the mechanism(s) which proteins exploit to gain heat resistance.
Guagliardi
A
A
Dipartimento di Chimica Organica e Biologica, Università di Napoli, Italy.
Martino
M
M
Iaccarino
I
I
De Rosa
M
M
Rossi
M
M
Bartolucci
S
S
D000426:Alcohol Dehydrogenase / Q000302:isolation & purification*; D004795:Enzyme Stability; D001411:Geobacillus stearothermophilus / Q000201:enzymology* / Q000254:growth & development; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D011489:Protein Denaturation; D013379:Substrate Specificity
D000426:Alcohol Dehydrogenase