CONNECTIVE-TISSUE MICROFIBRILS - ISOLATION AND CHARACTERIZATION OF 3 LARGE PEPSIN-RESISTANT DOMAINS OF FIBRILLIN

  • 15 December 1989
    • journal article
    • research article
    • Vol. 264  (35) , 21381-21385
Abstract
Human amnion was solubilized using pepsin and the digest supernatant screened for fragments of fibrillin with a previously characterized monoclonal antibody (Sakai, L. Y., Keene, D.R., and Engvall, E. (1986) J. Cell Biol. 103, 2499-2509). One fragment (PF1), with an apparent molecular weight of 94,000, was isolated and characterized. Two other fragments, PF2 and PF3, were isolated and shown to be fragments of fibrillin by preparing a monospecific antisera to PF2 and a monoclonal antibody to PF3. Immunoelectron microscopy and immunoblotting showed that both antibodies pictures of rotary-shadowed PF1 and PF2 showed them to be short rod-shaped molecules while PF3 has a crab-like appearance and seems to be an aggregate of several fibrillin chains fragments. Amino-terminal amino acid sequencing of PF1 and PF2 gave single unique sequences. Each of the three antibodies used was specific for one fragment and peptide mapping of PF1 and PF2 showed that there was no significant amino acids sequences overlap. Aggregates of PF3 are described which provided insight into the assembly and macromolecular structure of fibrillin in microfibrils.