Studies on the viscosity of solutions of bovine liver glutamate dehydrogenase and on related hydrodynamic models; effect of toluene on enzyme association
✍ Scribed by E. Reisler; H. Eisenberg
- Publisher
- Wiley (John Wiley & Sons)
- Year
- 1970
- Tongue
- English
- Weight
- 724 KB
- Volume
- 9
- Category
- Article
- ISSN
- 0006-3525
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✦ Synopsis
Abstract
The viscosity of bovine liver glutamate dehydrogenase solutions was studied at 10 and 20° C in 0.2.M sodium phosphate buffer at pH 7, in the concentration range 0.1–8 mg/ml. A method for the study of the viscosity of very dilute solutions of associating enzymes is described. It was found that the reduced specific viscosity η__~sp~/c__ of glutamate dehydrogenase continuously increases with increasing enzyme concentration, from about 4 ml/g at the lowest concentrations to about 16 ml/g at 8 mg/ml. In the presence of 10^−3^M GTP and 10^−3^M NADH the viscosity increase is much smaller and the results can be extrapolated to zero enzyme concentration to yield an intrinsic viscosity [η] = 3.2. The values of η__~sp~/c__ in phosphate buffer alone apparently extrapolate to the same value of [η], or to a value close to it. We also observe that, in the presence of toluene η__~sp~/c__ increases very much more with enzyme concentration: η__~sp~/c__ already equals 16 ml/g at a concentration of 0.75 mg/ml. These observations are in good agreement with the hypothesis that the active oligomer of glutamate dehydrogenase (MW = 312,000) associates with increasing enzyme concentration to form linear rodlike polymers of indefinite length. This association is strongly diminished by the addition of 10^−3^M GTP, 10^−3^M NADH. Toluene, on the other hand, promotes reversible association to linear polymers of very high molecular weight. The transverse and axial rotary frictional coefficients of macroscopic bodies, similar to a physical model for the structure of glutamate dehydrogenase recently advanced, were determined. Assuming that the viscosity of the model is equal to that of an ellipsoid of rotation with identical frictional coefficients, we calculate [η] = 3.26 ml/g according to Kuhn and 3.20 ml/g according to Simha, for the glutamate dehydrogenase oligomer, in good agreement with the result derived from the study of enzyme solutions.