Calcium determines the shape of fibrillin.
Velocity sedimentation experiments using authentic fibrillin-1 demonstrated sedimentation coefficients of s20,w0 = 5.1 +/- 0.1 in the Ca2+ form and s20,w0 = 6.2 +/- 0.1 in the Ca2+-free form. Calculations based on these results and the corresponding molecular mass predicted a shortening of fibrillin by approximately 25% and an increase in width of approximately 13-17% upon removal of Ca2+. These observations were confirmed by analysis of Ca2+-loaded and Ca2+-free rotary shadowed fibrillin molecules. Analysis of recombinant fibrillin-1 subdomain rF17, consisting primarily of an array of 12 Ca2+-binding epidermal growth factor (cbEGF)-like repeats, by analytical ultracentrifugation and rotary shadowing further confirmed Ca2+-dependent structural changes in the tertiary structure of fibrillin-1. Based on these results, the contribution of a single cbEGF-like repeat to the length of tandem arrays is predicted to be approximately 3 nm in the Ca2+ form. Ca2+-free forms demonstrated a decrease of 20-30% in length, indicating significant structural changes of these motifs when they occur in tandem. Circular dichroism measurements of rF17 in the presence and absence of Ca2+ indicated secondary structural changes within and adjacent to the interdomain regions that connect cbEGF-like repeats. The results presented here suggest a flexible structure for the Ca2+-free form of fibrillin which becomes stabilized, more extended, and rigid in the Ca2+ form.
Reinhardt
D P
DP
Shriners Hospital for Children, Portland, Oregon 97201, USA. dpr@shcc.org
Mechling
D E
DE
Boswell
B A
BA
Keene
D R
DR
Sakai
L Y
LY
Bächinger
H P
HP
D002118:Calcium / Q000737:chemistry*; D002942:Circular Dichroism; D016326:Extracellular Matrix Proteins / Q000737:chemistry; D000071838:Fibrillin-1; D000071837:Fibrillins; D006801:Humans; D008840:Microfilament Proteins / Q000737:chemistry / Q000648:ultrastructure*; D011994:Recombinant Proteins / Q000737:chemistry; D054392:Adipokines
D002118:Calcium; D016326:Extracellular Matrix Proteins; D000071838:Fibrillin-1; D000071837:Fibrillins; D008840:Microfilament Proteins; D011994:Recombinant Proteins; C000605722:FBN1 protein, human; D054392:Adipokines