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Extracellular Regulation of Bone Morphogenetic Protein Activity by the Microfibril Component Fibrillin-1.

Wohl AP, Troilo H, Collins RF, Baldock C, Sengle G - J. Biol. Chem. (2016)

Bottom Line: However, upon binding to fibrillin-1, the BMP-7 complex is rendered into a closed ring shape, which also confers latency to the growth factor, as demonstrated by bioactivity measurements.BMP-7 prodomain variants were used to map the critical epitopes for prodomain-growth factor and prodomain-prodomain binding.Together, these data show that upon prodomain binding to fibrillin-1, the BMP-7 complex undergoes a conformational change, which denies access of BMP receptors to the growth factor.

View Article: PubMed Central - PubMed

Affiliation: From the Center for Biochemistry, Medical Faculty, University of Cologne, Joseph-Stelzmann-Street 52, 50931 Cologne, Germany.

No MeSH data available.


Related in: MedlinePlus

Three-dimensional EM and solution SAXS models of BMP-7 PD-GF complex. Three-dimensional structure of BMP-7 complex was generated using TEM. A (top), representative electron micrograph of BMP-7 complex molecules (scale bar, 100 nm); bottom, 12 images selected from 140 class sum images of 9,000 particles that represent different views of BMP-7 complex (box size = 29.4 × 29.4 nm). B, class sum images were used to generate a three-dimensional TEM model of BMP-7 complex with 2-fold symmetry using angular reconstitution. C, superimposition of the BMP-9 complex structure (33) at 20 Å with the determined BMP-7 EM envelope suggests that the angle between the boomerang arms may be wider in BMP-9.
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Figure 2: Three-dimensional EM and solution SAXS models of BMP-7 PD-GF complex. Three-dimensional structure of BMP-7 complex was generated using TEM. A (top), representative electron micrograph of BMP-7 complex molecules (scale bar, 100 nm); bottom, 12 images selected from 140 class sum images of 9,000 particles that represent different views of BMP-7 complex (box size = 29.4 × 29.4 nm). B, class sum images were used to generate a three-dimensional TEM model of BMP-7 complex with 2-fold symmetry using angular reconstitution. C, superimposition of the BMP-9 complex structure (33) at 20 Å with the determined BMP-7 EM envelope suggests that the angle between the boomerang arms may be wider in BMP-9.

Mentions: BMP-7 complex alone or dialyzed together with the fibrillin-1 N terminus at a 1:4 molar ratio (total protein concentration 10–20 μg/ml) was negatively stained as described previously (24). Data were recorded at ×30,000 magnification on a Tecnai Biotwin microscope at 120 kV with a Gatan Orius CCD camera. Images were recorded with a 1-s exposure at defocus values of −0.5 to −1.6 μm at 3.5 Å/pixel (Fig. 2A). Single particle analysis was performed using EMAN2 (25). For BMP-7 complex, 9,000 particles were selected, and for BMP-7 dialyzed with fibrillin-1, 11,000 particles were selected, using a combination of manual and semiautomated picking. Following contrast transfer function correction, each data set was subjected to two-dimensional classification. For BMP-7 complex, a total of 140 projection averages were selected and used to generate an initial three-dimensional model with C2 symmetry. This model was used as a start seed for eight rounds of iterative refinement to produce a self-consistent three-dimensional structure. Resolution estimates were 35 Å using Fourier shell correlation with a cut-off value of 0.5. Modeling was performed using UCSF Chimera (26).


Extracellular Regulation of Bone Morphogenetic Protein Activity by the Microfibril Component Fibrillin-1.

Wohl AP, Troilo H, Collins RF, Baldock C, Sengle G - J. Biol. Chem. (2016)

Three-dimensional EM and solution SAXS models of BMP-7 PD-GF complex. Three-dimensional structure of BMP-7 complex was generated using TEM. A (top), representative electron micrograph of BMP-7 complex molecules (scale bar, 100 nm); bottom, 12 images selected from 140 class sum images of 9,000 particles that represent different views of BMP-7 complex (box size = 29.4 × 29.4 nm). B, class sum images were used to generate a three-dimensional TEM model of BMP-7 complex with 2-fold symmetry using angular reconstitution. C, superimposition of the BMP-9 complex structure (33) at 20 Å with the determined BMP-7 EM envelope suggests that the angle between the boomerang arms may be wider in BMP-9.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4933460&req=5

Figure 2: Three-dimensional EM and solution SAXS models of BMP-7 PD-GF complex. Three-dimensional structure of BMP-7 complex was generated using TEM. A (top), representative electron micrograph of BMP-7 complex molecules (scale bar, 100 nm); bottom, 12 images selected from 140 class sum images of 9,000 particles that represent different views of BMP-7 complex (box size = 29.4 × 29.4 nm). B, class sum images were used to generate a three-dimensional TEM model of BMP-7 complex with 2-fold symmetry using angular reconstitution. C, superimposition of the BMP-9 complex structure (33) at 20 Å with the determined BMP-7 EM envelope suggests that the angle between the boomerang arms may be wider in BMP-9.
Mentions: BMP-7 complex alone or dialyzed together with the fibrillin-1 N terminus at a 1:4 molar ratio (total protein concentration 10–20 μg/ml) was negatively stained as described previously (24). Data were recorded at ×30,000 magnification on a Tecnai Biotwin microscope at 120 kV with a Gatan Orius CCD camera. Images were recorded with a 1-s exposure at defocus values of −0.5 to −1.6 μm at 3.5 Å/pixel (Fig. 2A). Single particle analysis was performed using EMAN2 (25). For BMP-7 complex, 9,000 particles were selected, and for BMP-7 dialyzed with fibrillin-1, 11,000 particles were selected, using a combination of manual and semiautomated picking. Following contrast transfer function correction, each data set was subjected to two-dimensional classification. For BMP-7 complex, a total of 140 projection averages were selected and used to generate an initial three-dimensional model with C2 symmetry. This model was used as a start seed for eight rounds of iterative refinement to produce a self-consistent three-dimensional structure. Resolution estimates were 35 Å using Fourier shell correlation with a cut-off value of 0.5. Modeling was performed using UCSF Chimera (26).

Bottom Line: However, upon binding to fibrillin-1, the BMP-7 complex is rendered into a closed ring shape, which also confers latency to the growth factor, as demonstrated by bioactivity measurements.BMP-7 prodomain variants were used to map the critical epitopes for prodomain-growth factor and prodomain-prodomain binding.Together, these data show that upon prodomain binding to fibrillin-1, the BMP-7 complex undergoes a conformational change, which denies access of BMP receptors to the growth factor.

View Article: PubMed Central - PubMed

Affiliation: From the Center for Biochemistry, Medical Faculty, University of Cologne, Joseph-Stelzmann-Street 52, 50931 Cologne, Germany.

No MeSH data available.


Related in: MedlinePlus