Experimental and Computational Assessment of the Stability of Spermine Synthase Mutations Associated with Snyder–Robinson Syndrome

YERLİKAYA A.

Proceedings of the National Academy of Sciences India Section B - Biological Sciences, 2026 (Scopus)

Özet

Snyder–Robinson syndrome (SRS) is a rare X-linked recessive disorder caused by mutations in spermine synthase gene, resulting in altered spermidine/spermine ratios. Initial studies identified a G-to-A transition in intron 4 of SMS, producing a truncated protein and loss of enzymatic activity. Subsequent reports described missense mutations, including p.G56S and p.V132G, associated with variable cognitive and neurological outcomes. Here, we assessed the impact of these mutations on SMS stability and function using a reticulocyte lysate-based in vitro degradation assay and in silico analyses. Surprisingly, both wild-type and mutant SMS proteins remained stable, suggesting that cellular cofactors may be required for degradation. Notably, the G56S mutant showed markedly reduced SMS activity and was predominantly recovered from the urea-solubilized insoluble fraction, suggesting impaired protein solubility due to altered folding or structural assembly. DUET, SDM and mCSM analyses predicted both G56S and V132G substitutions to be locally destabilizing, with V132G exerting a stronger effect, whereas YASARA-based energy minimization indicated no significant global structural perturbation. Structural mapping showed that these residues are distal to the active site, implicating roles in dimerization or allosteric regulation rather than direct catalysis. Hydrogen-bonding analyses revealed altered local interactions upon mutation, supporting subtle structural changes that impair function. The alignment analysis indicated that Gly56 and Val132 are conserved among mammals but not in distant species. Collectively, these findings illuminate the molecular basis of SMS regulation and highlight the importance of distal, non-catalytic residues in modulating enzyme activity and stability, providing insights that may help future therapeutic strategies for SRS.