Divalent Cations and the Divergence of βγ-Crystallin Function

Abstract

The βγ-crystallin superfamily contains both β- and γ-crystallins of the vertebrate eye lens and the microbial calcium-binding proteins, all of which are characterized by a common double-Greek key domain structure. The vertebrate βγ-crystallins are long-lived structural proteins that refract light onto the retina. In contrast, the microbial βγ-crystallins bind calcium ions. The βγ-crystallin from the tunicate Ciona intestinalis (Ci-βγ) provides a potential link between these two functions. It binds calcium with high affinity and is found in a light-sensitive sensory organ that is highly enriched in metal ions. Thus, Ci-βγ is valuable for investigating the evolution of the βγ-crystallin fold away from calcium binding and toward stability in the apo form as part of the vertebrate lens. Here, we investigate the effect of Ca2+ and other divalent cations on the stability and aggregation propensity of Ci-βγ and human γS-crystallin (HγS). Beyond Ca2+, Ci-βγ is capable of coordinating Mg2+, Sr2+, Co2+, Mn2+, Ni2+, and Zn2+, although only Sr2+ is bound with comparable affinity to its preferred metal ion. The extent to which the tested divalent cations stabilize Ci-βγ structure correlates strongly with ionic radius. In contrast, none of the tested divalent cations improved the stability of HγS, and some of them induced aggregation. Zn2+, Ni2+, and Co2+ induce aggregation by interacting with cysteine residues, whereas Cu2+-mediated aggregation proceeds via a different binding site.

Aggregation pathways of human γ D crystallin induced by metal ions revealed by time dependent methods

“…Following studies have continue to investigate the aggregation process due to high concentration of metals (Domínguez-Calva et al, 2018;Ramkumar et al, 2018). In the case of γS-crystallin it was also found that some cations induce the aggregation formation by interacting with cysteine and methonine residues (Roskamp et al, 2019). Nevertheless, it is unlikely that proteins in the lens would be suddenly in contact with a high concentration of metal ions.…”

Cumulative deamidations of the major lens protein γS‐crystallin increase its aggregation during unfolding and oxidation

“…Deamidation and oxidation (in the presence of GSSG), as occurs in the aging lens, together enhance the aggregation of γS. There are seven cysteine residues in γS of which four are buried with low solvent‐accessible surface area (1–2%) and three have high accessibility, particularly C24 (21–77%) 36,39 . Of the buried cysteine residues, C36, C82, and C114 become more accessible in the deamidated mimics, while one of the accessible cysteine residues, C26, becomes less accessible 37 .…”

“…The modification levels in aged γS range from 25–60% at N14, 50–80% at N76, and 20–90% at N143 6,21,29,30,33,34 . These levels of deamidation correlate with their surface accessibility, that is, the labile polar Asn residues at positions 14, 76, and 143, have solvent exposures of ~55, 58, and 60%, respectively 35,36 . Their location is indicated on the solution NMR‐derived structure of γS in Figure 1 12 .…”

“…6,21,29,30,33,34 These levels of deamidation correlate with their surface accessibility, that is, the labile polar Asn residues at positions 14, 76, and 143, have solvent exposures of 55, 58, and 60%, respectively. 35,36 Their location is indicated on the solution NMR-derived structure of γS in Figure 1. 12 The level of γS with in vivo deamidations at all three sites is estimated to be about 30% based on our recently published data for deamidation at N14 (84%) and N76 (73%) and on previous reports for deamidation at N143 (about 50%).…”

Peer Review #2 of "Aggregation pathways of human γ D crystallin induced by metal ions revealed by time dependent methods (v0.1)"

“…Following studies have continue to investigate the aggregation process due to high concentration of metals (Domínguez-Calva et al, 2018;Ramkumar et al, 2018). In the case of γS-crystallin it was also found that some cations induce the agragation formation by interacting with cysteine and methonine residues (Roskamp et al, 2019). Nevertheless, it is unlikely that proteins in the lens would be suddenly in contact with a high concentration of metal ions.…”