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J. Cell Biol.,
Volume 140, Number 2, January 26, 1998 295-303

* Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri 63110 and The folding and trafficking of tropoelastin is
thought to be mediated by intracellular chaperones,
although the identity and role of any tropoelastin chaperone remain to be determined. To identify proteins
that are associated with tropoelastin intracellularly, bifunctional chemical cross-linkers were used to covalently stabilize interactions between tropoelastin and
associated proteins in the secretory pathway in intact
fetal bovine auricular chondrocytes. Immunoprecipitation of tropoelastin from cell lysates after cross-linking and analysis by SDS-PAGE showed the presence of
two proteins of ~74 kD (p74) and 78 kD (p78) that
coimmunoprecipitated with tropoelastin. Microsequencing of peptide fragments from a cyanogen bromide
digest of p78 identified this protein as BiP and sequence analysis identified p74 as the peptidyl-prolyl
cis-trans isomerase, FKPB65. The appearance of BiP
and FKBP65 in the immunoprecipitations could be
enhanced by the addition of brefeldin A (BFA) and
N-acetyl-leu-leu-norleucinal (ALLN) to the culture
medium for the final 4 h of labeling. Tropoelastin accumulates in the fused ER/Golgi compartment in the
presence of BFA if its degradation is inhibited by
ALLN (Davis, E.C., and R.P. Mecham. 1996. J. Biol.
Chem. 271:3787-3794). The use of BFA and other secretion-disrupting agents suggests that the association of tropoelastin with FKBP65 occurs in the ER. Results
from this study provide the first identification of a
ligand for an FKBP in the secretory pathway and suggest that the prolyl cis-trans isomerase activity of
FKBP65 may be important for the proper folding of the
proline-rich tropoelastin molecule before secretion.
Respiratory and Critical Care Division, Department of Medicine, Barnes-Jewish Hospital, St. Louis, Missouri 63110
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