Bromine is Essential to the Life in Assembling the Collagen IV

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(Last Updated On: April 23, 2017)
Role of Bromide in The Assembly of Collagen IV Scaffold

Bromide ions are necessary for the assembly of collagen IV scaffold that is an essential component of the basement membrane. Image: McCall et al. 2014

Recently, it has been found that bromine in the form of bromide ion constitutes one of the trace elements necessary for the survival of the living beings. It is required as a cofactor of the enzyme peroxidasin that catalyzes the formation of sulfilimine cross-links in the collagen fiber as a post-translational modification.

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Collagen IV Scaffold in the Basement Membrane

Collagen fibers are essential components of the basement membranes which perform essential roles in the morphogenesis, tissue repair after injury and many more. Collagen IV subunits in the basement membrane are cross-linked to each other by the sulfilimine bridge forming a scaffold that provides mechanical support and serves as a ligand for many  cell surface receptors like integrins.

Sulfilimine cross-link formation is catalyzed by a heme peroxidase, peroxidasin, found in the basement membrane. During the sulfilimine bridge formation, peroxidasin first forms hypobromous acid (HOBr) and hypochlorous acid (HOCl) from bromide (Br) and chloride (Cl) ions respectively which are then mediate the cross-link formation.

To account for the role of bromide ions, researchers grew Drosophila on a bromide deficient diet. Later on, they were found to be similar to that of the Drosophila with peroxidasin cross-link function mutation possessing developmental abnormalities like altered basement membrane and altered tissue morphogenesis. However, restoring the bromide in the diet, researchers observed that the Drosophila were able to grow into a normal phenotype.

Sulfilimine cross-links the two triple helices of the collagen IV scaffold through their NC1 domains forming a globular hexameric structure in which Met93 and Hyl211 are linked through the Sulfilimine Bridge.

Later on, researchers analyzed the effect of the halides including a pseudohalide thiocyanate in the formation of a sulfilimine cross-link in the cell culture. The result reveals that the thiocyanate SCN and I inhibit the reaction while Br enhances the cross-link formation.

Formation of Sulfilimine Crosslink in the Collagen IV Scaffold

Sulfilimine Crosslink in Collagen IV scaffold

Methionine Halosulfonium Intermediate may react with amine or water to form sulfilimine or sulfoxide respectively. Image: McCall et al. 2014

To investigate the selectivity of the Br- over the Cl ions, the researchers synthesized a cyclic sulfilimine compound, dehydromethionine wherein the methionine halosulfonium (HSI) reacts with either an amine or water to form a sulfilimine bridge or sulfoxide respectively. During this, the researchers made two hypotheses, (1) collagen IV sulfilimine bridge formation proceeds through the HSI of Met93 and (2) selectivity residues with bromosulfonium (S-Br) intermediate reacts with the ε-NH2 group of the Hyl211 more preferentially to form a sulfilimine bridge while chlorosulfonium (S-Cl) intermediate reacts with water more preferentially to form methionine sulfoxide with an inability to form cross-link.

To illustrate the biological effect of the bromide ions in the formation of sulfilimine cross-link in the collagen IV scaffold, the researchers grew Drosophila in a bromide deficient medium and compared their development with that of the Drosophila grown in the bromide supplemented medium. Larvae of the first generation grew in the bromide free media exhibited the delayed development and their continued growth in the same media led to their progeny (larvae of the second generation) significantly reduced survival.

In case of Drosophila with peroxidasin mutation, they have reduced the amount of collagen IV sulfilimine cross-links with perturbation of their midgut basement membrane. Therefore, researchers predicted that bromide depletion might least to the phenocopy of the peroxidasin mutants. The researchers compared the midgut of the larvae grown in the bromide free media with that of the two independent peroxidasin mutants and they found that the larvae grew in the bromide free diet displayed disrupted and split overall collagen IV scaffold in the same way as seen in the peroxidasin mutants.

Using the transmission electron microscopy, researchers were able to compare the ultrastructure of the larvae of both bromide free diet and peroxidasin mutant and what did they find is that basement membrane was irregular, thickened and occasionally diffused.

All these results led researchers to hypothesize that bromide ion and peroxidasin interact in vivo to strengthen the collagen IV scaffold that acts as a molecular corset to control the egg shape and its development.

Reference: Cell

Article doi: 10.1016/j.cell.2014.05.009

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