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DETERMINING THE PATHOGENICITY OF THE H307R VARIANT OF HUMAN BMP15 USING BIOINFORMATICS METHODS

Abstract

Ovarian dysgenesis (ODG) causes endocrinological imbalances and can, in its most-severe forms, result in complete female infertility. Defects in bone morphogenetic protein 15 (BMP15), which regulates folliculogenesis and oocyte maturation through activation of granulosa cells, are one of the sources of this condition. H307R is a human BMP15 variant of potential clinical relevance with respect to ODG presentation. This substitution, which replaces polar histidine (H) with charged arginine (R), is within the critical-to-function TGF-β2 domain, a region of confidence in existing protein structure models. To determine if this variant should be added to those known to cause ODG, we used in silico bioinformatics tools to assess sequence conservation and protein structure impacts. Histidine 307 was found to be invariant across 13 vertebrates, suggesting mutational changes to this site are not well tolerated. Multiple tools that assess homology, functional information and/or physicochemical similarity also agreed that H307R would likely induce deleterious effects on BMP15 tertiary structure. Molecular dynamic simulations (root-mean-square deviation [RMSD] and fluctuation [RMSF]) allow the comparison of wild-type and variant protein motion over time globally (RMSD) and by amino acid (RMSF). RMSD analysis revealed no generalizable change in motion of H307R relative to wild-type across the entirety of the simulation (20ns), but differences in BMP15 behavior were clear, and the average RMSD was reduced (8.78 vs 9.03). RMSF analysis revealed widespread reduction of H307R flexibility, with this reduction including the region of the TGF-β2 domain. Overall, these findings suggest that H307R is likely damaging to proper BMP15 function and might explain ODG symptoms in clinical cases which do not test positive for documented variants. Our results may therefore help lead to future diagnostic strategies for fertility-related conditions. However, validation of our outcome requires in vivo induction of ODG following the introduction of this variant in an appropriate model organism.

Acknowledgements

We thank the HudsonAlpha Institute for Biotechnology for providing a licensed copy of YASARA and running the MDS analyses of wildtype and variant proteins on their institutes server.

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