Metal ion dependency of microfibrils supports a rod-like conformation for fibrillin-1 calcium-binding epidermal growth factor-like domains

Cardy, C.M.; Handford, P.A.

Journal of Molecular Biology 276(5): 855-860


ISSN/ISBN: 0022-2836
PMID: 9566191
DOI: 10.1006/jmbi.1997.1593
Accession: 018003588

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The effects of the removal and replacement of divalent cations on the ultrastructure Of 10 to 12 nm fibrillin-1-containing microfibrils have been studied, in order to investigate the conformation of fibrillin-1 calcium binding epidermal growth factor-like (cbEGF-like) domains within the microfibril. The NMR structure of a covalently linked pair of cbEGF-like domains from fibrillin-1 recently identified a rigid rod-like conformation for the domain pair stabilized by interdomain calcium binding. This suggested that tandem arrays of fibrillin-1 cbEGF-like domains may adopt an extended conformation within a microfibril. If correct, then removal of bound calcium from fibrillin-1 would be expected to increase the flexibility of each cbEGF-like interdomain linkage, resulting in a decrease in the length of the interbead region of the microfibril (and thus a decrease in bead to bead periodicity), a concomitant increase in its diameter, and an overall increase in the flexibility of the microfibril. Our results show that removal of calcium by treatment with EGTA causes a large alteration of the microfibril structure, resulting in microfibrils with a reduced beaded periodicity, a disrupted interbead region and an increased flexibility. These effects are readily reversible by the readdition of calcium (in the form of CaCl2), but not by the addition of magnesium (MgCl2). This is consistent with conformational changes in cbEGF-like domains causing the major structural effects on the microfibril. These results provide the first direct experimental evidence to support an extended rod-like conformation for multiple tandem repeats of fibrillin-t cbEGF-like domains within the microfibril, as predicted by the NMR structure of an isolated fibrillin-I cbEGF-like domain pair.