INHIBITION OF THE PLASMA MEMBRANE CALCIUM PUMP BY AURINTRICARBOXYLIC ACID

SOUTO GUEVARA, CECILIA; FERREIRA GOMES, MARIELA; ROSSI JPFC; MANGIALAVORI IRENE

UNICAMP Convention Center, Campinas, SP

Congreso; 47th Annual Meeting of the Brazilian Biophysical Society; 2023

Plasma membrane calcium ATPases (PMCA) are high affinity calcium pumps that extrude calcium from the cytoplasm to the extracellular medium. There are four isoforms of PMCA, PMCA1 and PMCA4 are ubiquitous whereas PMCA2 and PMCA3 are restricted to certain tissues of the body (brain, ear, adrenal gland and cerebellum). They are involved in several physiological processes, as they intervene in the tight regulation of intracellular calcium, therefore playing a critical role in signaling pathways. However, it has been demonstrated that mutations in the genes that encode PMCAs are also involved in disease, such as cerebellar ataxia, infertility and primary hyperaldosteronism.Aurintricarboxylic acid (ATA) is a synthetic compound, produced by the condensation of salicylic acid and formaldehyde. It has been reported to inhibit several proteins, like bacterial phosphatases, viral integrases and helicases, including the inhibition of Spike from SARS CoV21. Mohamed et al2 proposed aurintricarboxylic acid as a specific inhibitor of PMCA4, but the mechanism of this inhibition remains unknown. We aim to study the interaction between ATA and PMCA (obtained from human erythrocytes, which is 90), in order to propose a binding site and a model of inhibition that could explain the results that we obtained:-ATA inhibits ATPase activity of PMCA with an apparent affinity of 70 nM. This inhibition is acompetitive with calcium and magnesium and not competitive with ATP.-ATA inhibits phosphatase activity of PMCA (carried in absence of calcium), with an apparent affinity of 90 nM.-ATA forms a complex with magnesium, detected by fluorescence. This complex would be the one that inhibits PMCA.-ATA acts as quencher of certain aromatic amino acids of PMCA and its apparent affinity for PMCA changes in presence of PMCA-ATP.This work was supported by Agencia Nacional de Promoción Científica y Tecnológica and Universidad de Buenos Aires.1 Minetti, C. A., Remeta, D. P., Hashimoto, K., Bonala, R., Chennamshetti, R., Yin, X., Garcia-Diaz, M., Grollman, A. P., Johnson, F., & Sidorenko, V. S. (2022). Characterization of Aurintricarboxylic Acid (ATA) Interactions with Plasma Transporter Protein and SARS-CoV-2 Viral Targets: Correlation of Functional Activity and Binding Energetics. Life, 12(6). https://doi.org/10.3390/life120608722Mohamed, T. M. A., Abou-Leisa, R., Baudoin, F., Stafford, N., Neyses, L., Cartwright, E. J., & Oceandy, D. (2013). Development and characterization of a novel fluorescent indicator protein PMCA4-GCaMP2 in cardiomyocytes. Journal of Molecular and Cellular Cardiology, 63, 57–68. https://doi.org/10.1016/j.yjmcc.2013.07.007