Self-assembly and supramolecular organization of EMILIN

The primary structure of human Elastin microfibril interface-located protein (EMILIN), an elastic fiber-as- sociated glycoprotein, consists of a globular C1q domain (gC1q) at the C terminus, a short collagenous stalk, a long region with a high potential for forming coiled-coil helices, and a cysteine-rich N-terminal sequence. It is not known whether the EMILIN gC1q domain is in- volved in the assembly process and in the supramolecu- lar organization as shown for the similar domain of col- lagen X. By employing the yeast two-hybrid system the EMILIN gC1q domains interacted with themselves, proving for the first time that this interaction occurs in vivo. The gC1q domain formed oligomers running as trimers in native gels that were less stable than the comparable trimers of the collagen X gC1q domain since they did not withstand heating. The collagenous domain was trypsin-resistant and migrated at a size correspond- ing to a triple helix under native conditions. In reducing agarose gels, EMILIN also migrated as a trimer, whereas under non-reducing conditions it formed polymers of many millions of daltons. A truncated fragment lacking gC1q and collagenous domains assembled to a much lesser extent, thus deducing that the C-terminal do- main(s) are essential for the formation of trimers that finally assemble into large EMILIN multimers.