HOMODIMERIC AND EXPANDED BEHAVIOR OF TRIMETHYLAMINE DEHYDROGENASE IN SOLUTION AT DIFFERENT TEMPERATURES

Earlier studies using crystallography have shown that trimethylamine d ehydrogenase (TMADH) from methylotropic bacteria exists as homodimers in the crystalline state. In this present hydrodynamic study we show t hat this is true also in dilute solution conditions and investigate th e degree of swelling or relaxation of the protein in solution. Analyti cal ultracentrifugation was used to determine the molar mass and to in vestigate whether the homodimeric nature of this molecule in crystal f orm - as visualized by x-ray crystallography - is reproduced in dilute solution at temperatures between 4 and 40 degrees C. The globular sol ution structure determined at 4 and 40 degrees C is in good agreement with crystallographic data although trimethylamine dehydrogenase was f ound to be either more asymmetric in solution - or highly hydrated -, a phenomenon found to increase with temperature. In agreement with the crystallographic structure, the enzyme sediments as a homodimer with a molar mass of(163,000+/-5,000) g/mol. The concentration dependence o f the sedimentation coefficient in the range of 0-1 mg/ml, indicates t hat no association or dissociation occurs. These findings are addition ally supported by sedimentation equilibrium data in the concentration range of 0 to 1.8 mg/ml. Finally, from the sedimentation coefficient d istribution at various temperatures, it was concluded that the enzyme is conformationally flexible and assumes an even more expanded structu re at higher temperatures which is in good agreement with the hydrodyn amic calculations performed.