Amidase activity and thermal stability of human thrombin.
Previous studies of amidase activity of human alpha-thrombin have yielded variable results and the decrease of this activity as a function of time and temperature has never been quantified. As this protease is an efficient tool in biochemistry and biotechnology thanks to its extreme selectivity, amidase activity and stability of thrombin were investigated with the synthetic substrate Tos-Gly-Pro-Arg-pNa. Enzyme activity as a function of temperature showed an optimum peak at 45 degrees C. The pH dependence of the activity showed a maximum around 9.5. The addition of NaCl promoted an increase of the activity. Stability of thrombin decreased rapidly when increasing the temperature from 25-45 degrees C and when diluting the enzyme. The presence of glycerol and ethylene glycol promoted a small increase of thrombin half life, whereas polyethylene glycol had a more pronounced positive effect even at very low concentrations.
Le Borgne
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Département de Génie Biochimique et Alimentaire, UA CNRS 544 INSA, Complexe Scientifique de Rangueil, Toulouse, France.
Graber
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D000581:Amidohydrolases / Q000378:metabolism*; D000595:Amino Acid Sequence; D004795:Enzyme Stability; D019855:Ethylene Glycol; D005026:Ethylene Glycols / Q000494:pharmacology; D005990:Glycerol / Q000494:pharmacology; D006358:Hot Temperature; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D012965:Sodium Chloride / Q000494:pharmacology; D013917:Thrombin / Q000378:metabolism*
D005026:Ethylene Glycols; D012965:Sodium Chloride; D013917:Thrombin; D000581:Amidohydrolases; C019665:amidase; D019855:Ethylene Glycol; D005990:Glycerol