LTBP-2 competes with tropoelastin for binding to fibulin-5 and heparin, and is a negative modulator of elastinogenesis
| Autor: | Mark Gibson, Mahroo K. Parsi, Clementine Menz, Mohamed Arshad Mohamed Sideek |
|---|---|
| Rok vydání: | 2014 |
| Předmět: |
Fibrillin-1
Elastic fiber assembly Perlecan Fibrillins Mice 03 medical and health sciences chemistry.chemical_compound 0302 clinical medicine Tropoelastin medicine Animals Humans Binding site Molecular Biology Cells Cultured 030304 developmental biology Extracellular Matrix Proteins 0303 health sciences integumentary system biology Heparin Microfilament Proteins Heparan sulfate Fibroblasts Elastic Tissue Fibulin Latent TGF-beta Binding Proteins chemistry Biochemistry 030220 oncology & carcinogenesis biology.protein Biophysics Heparitin Sulfate Elastin Protein Binding medicine.drug |
| Zdroj: | Matrix Biology. 34:114-123 |
| ISSN: | 0945-053X |
| DOI: | 10.1016/j.matbio.2013.10.007 |
| Popis: | Latent transforming growth factor-beta-1 binding protein-2 (LTBP-2) is a protein of ill-defined function associated with elastic fibers during elastinogenesis. Although LTBP-2 binds fibrillin-1, fibulin-5, and heparin/heparan sulfate, molecules critical for normal elastic fiber assembly, it does not interact directly with elastin or its precursor, tropoelastin. We investigated the modulating effect of LTBP-2 on two key interactions of tropoelastin during elastinogenesis a) with fibulin-5 and b) with heparan sulfate (using heparin). Firstly, using solid phase assays we showed that LTBP-2 bound fibulin-5 (Kd=26.47±5.68 nM) with an affinity similar to that of the tropoelastin-fibulin-5 interaction (Kd=24.66±5.64 nM). Then using a competitive binding assay we showed that LTBP-2 inhibited the tropoelastin-fibulin-5 interaction in a dose dependent manner with almost complete inhibition obtained with 5-fold molar excess of LTBP-2. Interestingly, a fragment of LTBP-2 containing the fibulin-5 binding sequence only partially inhibited the tropoelasin-fibulin-5 interaction suggesting that LTBP-2 was directly blocking only the C-terminal tropoelastin binding site on fibulin-5 and indirectly blocking tropoelastin binding to the N-terminal region. In parallel experiments heparin was shown to have minor inhibitory effects on fibulin-5 interactions with tropoelastin and LTBP-2. However, LTBP-2 was shown to significantly inhibit the binding of heparin to tropoelastin with 50% inhibition achieved with 10 fold molar excess of LTBP-2. Confocal microscopy of fibroblast matrix showed strong co-distribution of LTBP-2 with fibulin-5 and fibrillin-1 and partial co-distribution with heparan sulfate proteoglycans, perlecan and syndecan-4. Also addition of exogenous LTBP-2 to ear cartilage chondrocyte cultures blocked elastinogenesis in a concentration-dependent manner. Overall the results indicate that LTBP-2 may have a negative regulatory role during elastic fiber assembly, perhaps in displacing elastin microassemblies from complexes with fibulin-5 and/or cell surface heparan sulfate proteoglycans. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|
Full Text from ScienceDirect