The Subunit Composition and Sedimentation Properties of Human C1

Abstract

Abstract An equimolar ratio of C1q, C1r, and C1s was found in normal human serum by radial immunodiffusion when 410,000, 190,000, and 174,000 daltons were used as their respective m.w. These values for C1r and C1s represent the sum of the weights of the two noncovalently linked subunits of C1r and the sum of the two calcium-linked subunits of C1s. Up to 95% of the serum C1 hemolytic activity applied to sucrose gradients formed in buffers of physiologic ionic strength was recovered from the gradients in a single peak which sedimented with a rate of 16S. After pooling, dialysis, and concentration, this peak was analyzed by single radial immunodiffusion and was found to contain equimolar concentrations of C1q, C1r, and C1s. The specific activity of this peak of C1, when expressed as effective molecules per microgram of C1q, C1r, and C1s was identical to that found in serum. C1 reassembled from C1q, C1r, and C1s sedimented as a 15 to 16S macromolecule and also contained equimolar concentrations of C1q, C1r, and C1s. With dilution of serum before centrifugation, the recovery of C1 from the gradients dropped dramatically and only 18% of the C1 activity was recovered from a 1/10 dilution. Under these conditions, minor peaks of C1 activity representing only a few percent of C1 activity became evident at 12S and 19S. Under low ionic strength conditions C1 aggregated. C1t antigen was found to behave independently from C1 in these studies. The number of moles of C1t in serum varied from one-third to two-thirds of that observed for C1q, C1r, and C1s. In sucrose gradients all of the C1t sedimented separately from C1 with a rate of <13S. Isolated C1t did not substitute for any C1 subunit or enhance C1 activity or the sedimentation rate of C1. Thus, no role for C1t in C1 activity or structure was evident in these studies. This work indicates that C1 is composed of equimolar concentrations of C1q, C1r, and C1s assembled in a macromolecule with a m.w. of 774,000 daltons which sediments in sucrose gradients with a rate of 16S.