The Somatomedin C Binding Protein: Evidence for a Heterologous Subunit Structure*

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Abstract
The binding of somatomedin C to serum proteins was investigated using Sephadex G-200 chromatography at pH 7.40. Under these conditions, immunoreactive somatomedin C in whole serum has a Stokes' radius of 43 Å (Kd = 0.25). After ammonium sulfate precipitation and DEAE Sephadex chromatography, three protein peaks were obtained. The second peak contained the majority of the immunoreactive somatomedin C, but the somatomedin in this fraction has Stokes' radii of 36 and 14 Å (Kd = 0.35 and 0.75, respectively). Recombining this fraction with the DEAE Sephadex peak 3 fraction resulted in the reappearance of the 43 Å (Kd = 0.25) species. Chromatography of the 43 Å somatomedin C species on Sephadex G-50 in 0.1 M acetic acid-0.15 M NaCl dissociated somatomedin C from its binding protein. The binding protein was recovered in the void volume fractions of the G-50 column. This acid-treated binding protein had a Stokes' radius of 36 Å (Kd = 0.35), as determined by the binding of [l25I]somatomedin C at pH 7.40. When the acid-treated binding protein was incubated with the DEAE Sephadex peak 3 fraction, the binding of [125I]somatomedin C was shifted to the 43 Å (Kd = 0.25) region. The DEAE Sephadex peak 3 material is destroyed by brief exposure to low pH and it is absent from the serum of GHdeficient patients. The 36 Å binding species is also absent from the serum of GH-deficient patients. Both the DEAE Sephadex peak 3 material and the 36 Å binding species are present in the serum of GH-deficient patients given parenteral GH. Serum also contains a second acid-stable protein which binds somatomedin C. This protein has a Stokes' radius of 28.5 Å (Kd = 0.50) and it is present in the serum of both normal and GHdeficient patients. The protein does not recombine with the DEAE Sephadex peak 3 protein. These results suggest that the somatomedin C binding activity in whole serum is composed of at least two subunits, both of which are under GH control. A second, smaller, non-GH-dependent binding species is also present in serum, but it does not appear to be a major factor in transporting the endogenous somatomedin C activity.