The fourth component of human complement is an essential part of the classical and lectin pathways performing multifunctional roles in both host defense and immune regulation. C4 is the most polymorphic member of the complement proteins, and complete deficiency is strongly associated with autoimmune disease, especially, systemic lupus erythematosus (SLE). Of the two C4 genes C4A, but not C4B, null alleles have been implicated as important independent disease susceptibility genes occurring in more than half of SLE patients. Whether and how this deficiency contributes to the development or pathology remains unclear. We do know that activation of C4 by C1s cleaves the thioester bond, thus inducing a conformational change that exposes numerous ligand-binding sites involved in functional activity. Structural comparison, among many other tools, plays an important role in predicting function. In this report, the tertiary structures of C4A and C4B were compared using near and far-UV circular dichroism, ANS fluorescence, site-specific monoclonal antibodies and isoelectric focusing. Negligible differences in the native proteins were found. However, the activated proteins were dissimilar in secondary and tertiary structure that was accompanied by significant differences in charge distribution and surface hydrophobicity. These conformational differences, together with known acceptor preferences, have functional implications for the association between C4A null alleles and SLE.