The Highly Conservative Cysteine of Oncomodulin as a Feasible Redox Sensor.

Alisa A Vologzhannikova, Polina A Khorn, Marina P Shevelyova, Alexei S Kazakov, Victor I Emelyanenko, Eugene A Permyakov, Sergei E Permyakov

Biomolecules 2021 Jan 06

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Oncomodulin (Ocm), or parvalbumin β, is an 11-12 kDa Ca2+-binding protein found inside and outside of vertebrate cells, which regulates numerous processes via poorly understood mechanisms. Ocm consists of two active Ca2+-specific domains of the EF-hand type ("helix-loop-helix" motif), covered by an EF-hand domain with inactive EF-hand loop, which contains a highly conservative cysteine with unknown function. In this study, we have explored peculiarities of the microenvironment of the conservative Cys18 of recombinant rat Ocm (rWT Ocm), redox properties of this residue, and structural/functional sensitivity of rWT Ocm to the homologous C18S substitution. We have found that pKa of the Cys18 thiol lays beyond the physiological pH range. The measurement of redox dependence of rWT Ocm thiol-disulfide equilibrium (glutathione redox pair) showed that redox potential of Cys18 for the metal-free and Ca2+-loaded protein is of -168 mV and -176 mV, respectively. Therefore, the conservative thiol of rWT Ocm is prone to disulfide dimerization under physiological redox conditions. The C18S substitution drastically reduces α-helices content of the metal-free and Mg2+-bound Ocm, increases solvent accessibility of its hydrophobic residues, eliminates the cooperative thermal transition in the apo-protein, suppresses Ca2+/Mg2+ affinity of the EF site, and accelerates Ca2+ dissociation from Ocm. The distinct structural and functional consequences of the minor structural modification of Cys18 indicate its possible redox sensory function. Since some other EF-hand proteins also contain a conservative redox-sensitive cysteine located in an inactive EF-hand loop, it is reasonable to suggest that in the course of evolution, some of the EF-hands attained redox sensitivity at the expense of the loss of their Ca2+ affinity.

Citation

Alisa A Vologzhannikova, Polina A Khorn, Marina P Shevelyova, Alexei S Kazakov, Victor I Emelyanenko, Eugene A Permyakov, Sergei E Permyakov. The Highly Conservative Cysteine of Oncomodulin as a Feasible Redox Sensor. Biomolecules. 2021 Jan 06;11(1)