Some physical properties of purified fraction I protein from spinach chloroplasts

Abstract

Fraction I protein, from spinach chloroplasts and homogeneous by electrophoretic and sedimentation velocity criteria, had an uncorrected specific refractive increment of 1.69 × 10 −4 ml/mg protein at 10.0 ° and 546 mμ. Its specific extinction coefficient was 1.82 at 1 mg/ml in pH 8.0, 0.001 m Tris-chloride for a 1-cm light path. Moving boundary electrophoresis of this protein at about 4 ° yielded mobilities from −3.2 to −6.2 × 10 −5 cm 2 volt −1 sec −1 at pH 6.3–9.5, which, on extrapolation, indicated an isoelectric point around pH 4.3–5.5. The sedimentation coefficient decreased with increasing protein concentration above 0.3 mg/ml and was 18.7S when extrapolated to zero concentration. At less than 0.3 mg/ml, the sedimentation coefficient decreased with decreasing concentration, suggesting the dissociation of the protein. The apparent diffusion coefficient was 3.01 × 10 −7 cm 2 sec −1, which when combined with the extrapolated sedimentation coefficient gave a molecular weight of 559,000. Sedimentation equilibrium experiments gave a limiting molecular weight at the meniscus of 475,000 ± 11,000 and suggested the presence of a 545,000 molecular weight component. An attempt was made to explain the 17% discrepancy between the molecular weights from sedimentation velocity runs and that from sedimentation equilibrium runs