Open AccessDetermination of Molecular Weight and Molecular Structurof Rat‐Liver Pyruvate Carboxylase
Author(s) -
GOTTSCHALK EllenMarie,
MAYER Frank,
KLOSTERMANN Alice,
SEUBERT Werner
Publication year - 1976
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1976.tb10317.x
Subject(s) - tetramer , ultracentrifuge , pyruvate carboxylase , sedimentation coefficient , molecular mass , chemistry , electron microscope , sedimentation equilibrium , protein subunit , biochemistry , enzyme , crystallography , physics , optics , gene
The molecular weight of pyruvate carboxylase isolated from pigeon and rat liver mitochondria was examined using analytical ultracentrifugation and electron microscopy. The enzyme molecule appeared as a tetramer with the four subunits arranged at the corners of a square. Sedimentation studies in the analytical ultracentrifuge, extrapolated to infinite dilution, showed the tetramer to have a molecular weight M r c = 0 of 280000 and an s 20,w 0 of 12.7 S. The tetramer could be dissociated into trimers and dimers of lower specific enzymic activity by storage at 4°C or incubation at ‐20°C at low protein concentrations. The isolated trimers and dimers had a molecular weight M r c = 0 of 210000 and 140000, respectively, and an s 20,w 0 of 10.85 S and 7.55 S, respectively. Incubation with 2 M urea at 20°C yielded enzymically inactive subunits (M r c = 0 = 70000; s 20,w 0 = 4.95 S). The molecular weights (for pyruvate carboxylase and its subunits), as calculated from the subunit diameter observed in the electron microscope, were consistent with the values obtained from sedimentation studies.