Clinical Chemistry
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Clinical Chemistry 27: 213-219, 1981;
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Clinical Chemistry, Vol 27, 213-219, Copyright © 1981 by American Association for Clinical Chemistry

Effects of temperature on the steady-state kinetics and measurement of aspartate aminotransferases

R Rej and RE Vanderlinde

We examined the effects of temperature on the activity and steady-state kinetics of aspartate aminotransferase (EC 2.6.1.1), using purified human soluble (s-AspAT) and mitochondrial (m-AspAT) isoenzymes, human serum, and porcine s-AspAT. All enzymes obeyed similar linear Arrhenius relationships over the range 20-40 degrees C. Apparent energies of activation (52.3 kJ.mol-1) and ratios of activity between 30 and 37 degrees C (0.626) were identical for the human s- and m-AspAT. This ratio was 0.623 (SEM 0.004) for human sera; deviation from the predicted ratio by individual sera was within analytical error. Similar activity/temperature relationships were observed for porcine s-AspAT. The use of factors to convert AspAT activities at 30 and 37 degrees C influenced neither precision of measurement of frequency distributions of results. The apparent Michaelis constants for the human isoenzymes increased with temperature. The least-influenced Km was for 2- oxoglutarate and s-AspAT: K2-oxoglutarate was 0.24 mmol.L-1 at 25 degrees C and 0.29 mmol.L-1 at 37 degrees C; apparent enthalpy change for substrate binding (delta HS) was 12.1 kJ.mol-1. The largest variation was for 2-oxoglutarate and m-AspAT: K2-oxoglutarate was 0.46 mmol.L-1 at 25 degrees C and 1.02 mmol.L-1 at 37 degrees C; delta HS was 50.8 kJ.mol-1. Incubation of the human isoenzymes with substrate mixture (without 2-oxoglutarate) at 23 and 37 degrees C did not affect activity during 60 min if tris(hydroxymethyl)aminomethane buffer was used. When the isoenzymes were diluted to 10 nmol-L-1 (about 200 U.L-1) in buffer alone and incubated at 50 degrees C, m-AspAT activity was decreased by 20% after 120 min; the cytoplasmic enzyme was unaffected.


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 Clin. Chem.Home page
L. F. Perez and J. C. Tutor
Assay of ß-N-acetylhexosaminidase isoenzymes in different biological specimens by means of determination of their activation energies
Clin. Chem., February 1, 1998; 44(2): 226 - 231.
[Abstract] [Full Text] [PDF]


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Copyright © 1981 by the American Association for Clinical Chemistry.