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Structural effects of fibulin 5 missense mutations associated with age-related macular degeneration and cutis laxa

Structural effects of fibulin 5 missense mutations associated with age-related macular degeneration and cutis laxa
Structural effects of fibulin 5 missense mutations associated with age-related macular degeneration and cutis laxa
PURPOSE: AMD has a complex etiology with environmental and genetic risk factors. Ten fibulin 5 sequence variants have been associated with AMD and two other fibulin 5 mutations cause autosomal-recessive cutis laxa. Fibulin 5 is a 52-kDa calcium-binding epidermal growth factor (cbEGF)-rich extracellular matrix protein that is essential for the formation of elastic tissues. Biophysical techniques were used to detect structural changes in the fibulin 5 mutants and to determine whether changes are predictive of pathogenicity.

METHODS: Native PAGE, nonreduced SDS-PAGE, size-exclusion column multiangle laser light scattering, sedimentation velocity, and circular dichroism (CD) were used to investigate the mobility, hydrodynamic radii, folding, and oligomeric states of the fibulin 5 mutants in the absence and presence of Ca(2+).

RESULTS: CD showed that all mutants are folded, although perturbations to secondary structure contents were detected. Both cutis laxa mutants increased dimerization. Most other mutants slightly increased self-association in the absence of Ca(2+) but this was also demonstrated by G202R, a polymorphism detected in a control individual. The AMD-associated mutant G412E showed lower-than-expected mobility during native-PAGE, the largest hydrodynamic radius for the monomer form and the highest levels of aggregation in both the absence and presence of Ca(2+).

CONCLUSIONS: The results identified structural differences for the disease-causing cutis laxa mutants and for one AMD variant (G412E), suggesting that this may also be pathogenic. Although the other AMD-associated mutants showed no gross structural differences, they cannot be excluded as pathogenic by differences outside the scope of this study-for example, disruption of heterointeractions.
0146-0404
2356-2362
Jones, R.P.O.
6fda8b1c-c504-483c-b657-f0ab87ee5cac
Ridley, C.
24fdbfa1-cedd-4016-a082-9c3f8c44041d
Jowitt, T.A.
ac97682b-9ec6-40b4-a9ab-481f1b29f240
Wang, M.C.
534308cb-ce36-47ed-a2ef-34a5ea4dccd2
Howard, M.
c0eb3016-5559-4d27-b7a2-ed81e2098300
Bobola, N.
cb7767c5-e10e-4993-a14e-5b0746eb3592
Wang, T.
6ebae192-c5d9-4506-8780-82db62caa1ca
Bishop, P.N.
3ef01b90-9eec-43d2-b160-0fde79f8bed2
Kielty, C.M.
414fa49a-9854-4736-8cab-76c051560bfb
Baldock, C.
7cb9e2ca-c282-4e89-b0d6-493246661b9d
Lotery, A.J.
5ecc2d2d-d0b4-468f-ad2c-df7156f8e514
Trump, D.
3bf4960e-1b72-4a77-929e-7179f9d991b5
Jones, R.P.O.
6fda8b1c-c504-483c-b657-f0ab87ee5cac
Ridley, C.
24fdbfa1-cedd-4016-a082-9c3f8c44041d
Jowitt, T.A.
ac97682b-9ec6-40b4-a9ab-481f1b29f240
Wang, M.C.
534308cb-ce36-47ed-a2ef-34a5ea4dccd2
Howard, M.
c0eb3016-5559-4d27-b7a2-ed81e2098300
Bobola, N.
cb7767c5-e10e-4993-a14e-5b0746eb3592
Wang, T.
6ebae192-c5d9-4506-8780-82db62caa1ca
Bishop, P.N.
3ef01b90-9eec-43d2-b160-0fde79f8bed2
Kielty, C.M.
414fa49a-9854-4736-8cab-76c051560bfb
Baldock, C.
7cb9e2ca-c282-4e89-b0d6-493246661b9d
Lotery, A.J.
5ecc2d2d-d0b4-468f-ad2c-df7156f8e514
Trump, D.
3bf4960e-1b72-4a77-929e-7179f9d991b5

Jones, R.P.O., Ridley, C., Jowitt, T.A., Wang, M.C., Howard, M., Bobola, N., Wang, T., Bishop, P.N., Kielty, C.M., Baldock, C., Lotery, A.J. and Trump, D. (2010) Structural effects of fibulin 5 missense mutations associated with age-related macular degeneration and cutis laxa. Investigative Ophthalmology & Visual Science, 51 (5), 2356-2362. (doi:10.1167/iovs.09-4620). (PMID:20007835)

Record type: Article

Abstract

PURPOSE: AMD has a complex etiology with environmental and genetic risk factors. Ten fibulin 5 sequence variants have been associated with AMD and two other fibulin 5 mutations cause autosomal-recessive cutis laxa. Fibulin 5 is a 52-kDa calcium-binding epidermal growth factor (cbEGF)-rich extracellular matrix protein that is essential for the formation of elastic tissues. Biophysical techniques were used to detect structural changes in the fibulin 5 mutants and to determine whether changes are predictive of pathogenicity.

METHODS: Native PAGE, nonreduced SDS-PAGE, size-exclusion column multiangle laser light scattering, sedimentation velocity, and circular dichroism (CD) were used to investigate the mobility, hydrodynamic radii, folding, and oligomeric states of the fibulin 5 mutants in the absence and presence of Ca(2+).

RESULTS: CD showed that all mutants are folded, although perturbations to secondary structure contents were detected. Both cutis laxa mutants increased dimerization. Most other mutants slightly increased self-association in the absence of Ca(2+) but this was also demonstrated by G202R, a polymorphism detected in a control individual. The AMD-associated mutant G412E showed lower-than-expected mobility during native-PAGE, the largest hydrodynamic radius for the monomer form and the highest levels of aggregation in both the absence and presence of Ca(2+).

CONCLUSIONS: The results identified structural differences for the disease-causing cutis laxa mutants and for one AMD variant (G412E), suggesting that this may also be pathogenic. Although the other AMD-associated mutants showed no gross structural differences, they cannot be excluded as pathogenic by differences outside the scope of this study-for example, disruption of heterointeractions.

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Published date: May 2010
Organisations: Clinical & Experimental Sciences

Identifiers

Local EPrints ID: 359179
URI: http://eprints.soton.ac.uk/id/eprint/359179
ISSN: 0146-0404
PURE UUID: ad5ac46c-6342-4544-9bbe-b62a11517f2c
ORCID for A.J. Lotery: ORCID iD orcid.org/0000-0001-5541-4305

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Date deposited: 23 Oct 2013 13:02
Last modified: 15 Mar 2024 03:16

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Contributors

Author: R.P.O. Jones
Author: C. Ridley
Author: T.A. Jowitt
Author: M.C. Wang
Author: M. Howard
Author: N. Bobola
Author: T. Wang
Author: P.N. Bishop
Author: C.M. Kielty
Author: C. Baldock
Author: A.J. Lotery ORCID iD
Author: D. Trump

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