Initial steps in assembly of microfibrils: Formation of disulfide-cross- linked multimers containing fibrillin-1

Dieter P. Reinhard, Jay E. Gambee, Robert N. Ono, Hans Peter Bächinger, Lynn Sakai

Research output: Contribution to journalArticle

61 Citations (Scopus)

Abstract

Fibrillins are the major constituents of extracellular microfibrils. How fibrillin molecules assemble into microfibrils is not known. Sequential extractions and pulse-chase labeling of organ cultures of embryonic chick aortae revealed rapid formation of disulfide-crosslinked aggregates containing fibrillin-1. These results demonstrated that intermolecular disulfide bond formation is an initial step in the assembly process. To identify free cysteine residues available for intermolecular cross-linking, small recombinant peptides of fibrillin-1 harboring candidate cysteine residues were analyzed. Results revealed that the first four cysteine residues in the unique N terminus form intramolecular disulfide bonds. One cysteine residue (Cys204) in the first hybrid domain of fibrillin-1 was found to occur as a free thiol and is therefore a good candidate for intermolecular disulfide bonding in initial steps of the assembly process. Furthermore, evidence indicated that the comparable cysteine residue in fibrillin-2 (Cys233) also occurs as a free thiol. These free cysteine residues in fibrillins are readily available for intermolecular disulfide bond formation, as determined by reaction with Ellman's reagent. In addition to these major results, the cleavage site of the fibrillin-1 signal peptide and the N-terminal sequence of monomeric authentic fibrillin-1 from conditioned fibroblast medium were determined.

Original languageEnglish (US)
Pages (from-to)2205-2210
Number of pages6
JournalJournal of Biological Chemistry
Volume275
Issue number3
DOIs
StatePublished - Jan 21 2000

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Microfibrils
Disulfides
Cysteine
Sulfhydryl Compounds
Dithionitrobenzoic Acid
Organ Culture Techniques
Fibroblasts
Conditioned Culture Medium
Protein Sorting Signals
Labeling
Aorta
Fibrillin-1
Peptides
Molecules
Fibrillins

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Initial steps in assembly of microfibrils : Formation of disulfide-cross- linked multimers containing fibrillin-1. / Reinhard, Dieter P.; Gambee, Jay E.; Ono, Robert N.; Bächinger, Hans Peter; Sakai, Lynn.

In: Journal of Biological Chemistry, Vol. 275, No. 3, 21.01.2000, p. 2205-2210.

Research output: Contribution to journalArticle

Reinhard, Dieter P. ; Gambee, Jay E. ; Ono, Robert N. ; Bächinger, Hans Peter ; Sakai, Lynn. / Initial steps in assembly of microfibrils : Formation of disulfide-cross- linked multimers containing fibrillin-1. In: Journal of Biological Chemistry. 2000 ; Vol. 275, No. 3. pp. 2205-2210.
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