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Here, we characterized the structural and biophysical consequences of cumulative Asn to Asp changes in \u03b3D\u2010crystallin. Using NMR spectroscopy, we demonstrate that N\u2010 or C\u2010terminal domain\u2010confined or fully Asn to Asp changed \u03b3D\u2010crystallin exhibits essentially the same <jats:sup>1<\/jats:sup>H\u2010<jats:sup>15<\/jats:sup>N HSQC spectrum as the wild\u2010type protein, implying that the overall structure is retained. Only a very small thermodynamic destabilization for the overall Asn to Asp \u03b3D\u2010crystallin variants was noted by chaotropic unfolding, and assessment of the colloidal stability, by measuring diffusion interaction parameters, yielded no substantive differences in association propensities. Furthermore, using molecular dynamics simulations, no significant changes in dynamics for proteins with Asn to Asp or iso\u2010Asp changes were detected. Our combined results demonstrate that substitution of all Asn by Asp residues, reflecting an extreme case of deamidation, did not affect the structure and biophysical properties of \u03b3D\u2010crystallin. This suggests that these changes alone cannot be the major determinant in driving cataract formation.<\/jats:p>","DOI":"10.1002\/pro.5120","type":"journal-article","created":{"date-parts":[[2024,7,18]],"date-time":"2024-07-18T09:47:03Z","timestamp":1721296023000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["Cumulative asparagine to aspartate deamidation fails to perturb <scp>\u03b3D<\/scp>\u2010crystallin structure and stability"],"prefix":"10.1002","volume":"33","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-9796-5719","authenticated-orcid":false,"given":"Alex J.","family":"Guseman","sequence":"first","affiliation":[{"name":"Department of Structural Biology University of Pittsburgh School of Medicine  Pittsburgh Pennsylvania USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jeremy J.","family":"Gonz\u00e1lez","sequence":"additional","affiliation":[{"name":"Department of Structural Biology 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