A finely tuned interplay between calcium binding, ionic strength and pH modulates conformational and oligomerization equilibria in the Respiratory Syncytial Virus Matrix (M) protein.

Autor: Esperante SA; Protein Structure-Function and Engineering Laboratory, Fundación Instituto Leloir and IIBBA-CONICET, CABA, Argentina. Electronic address: sesperante@infant.org.ar., Alvarez-Paggi D; INFANT Foundation and CONICET, Argentina. Electronic address: dalvarezpaggi@infant.org.ar., Salgueiro M; Protein Structure-Function and Engineering Laboratory, Fundación Instituto Leloir and IIBBA-CONICET, CABA, Argentina. Electronic address: marianosalgueirounq@gmail.com., Desimone MF; Universidad de Buenos Aires, Instituto de Química y Metabolismo del Fármaco (IQUIMEFA, CONICET-UBA), Facultad de Farmacia y Bioquímica, Cátedra de Química Analítica Instrumental, Buenos Aires, Argentina. Electronic address: desimone@ffyb.uba.ar., de Oliveira GAP; Programa de Biologia Estrutural, Instituto de Bioquímica Médica Leopoldo de Meis, Instituto, Nacional de Ciência e Tecnologia de Biologia Estrutural e Bioimagem, Centro Nacional de, Ressonância Magnêtica Nuclear Jiri Jonas, Argentina. Electronic address: gaugusto@bioqmed.ufrj.br., Arán M; NMR Laboratory, Fundación Instituto Leloir and IIBBA-CONICET, CABA, Argentina. Electronic address: maran@leloir.org.ar., García-Pardo J; IBB - Institut de Biotecnologia i Biomedicina - Universitat Autonoma de Barcelona, Barcelona, Spain. Electronic address: javiergarciapardo@msn.com., Aptekmann AA; Departamento de Química Biológica, IQUIBICEN-CONICET, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, Argentina. Electronic address: aaptekmann@qb.fcen.uba.ar., Ventura S; IBB - Institut de Biotecnologia i Biomedicina - Universitat Autonoma de Barcelona, Barcelona, Spain. Electronic address: salvador.ventura@uab.cat., Alonso LG; Instituto de Nanobiotecnología (NANOBIOTEC), CONICET-Universidad de Buenos Aires, Argentina. Electronic address: leonardoe7@gmail.com., de Prat-Gay G; Protein Structure-Function and Engineering Laboratory, Fundación Instituto Leloir and IIBBA-CONICET, CABA, Argentina. Electronic address: gpg@leloir.org.ar.
Jazyk: angličtina
Zdroj: Archives of biochemistry and biophysics [Arch Biochem Biophys] 2022 Nov 30; Vol. 731, pp. 109424. Date of Electronic Publication: 2022 Oct 08.
DOI: 10.1016/j.abb.2022.109424
Abstrakt: As in most enveloped RNA viruses, the Respiratory Syncytial Virus Matrix (RSV-M) protein plays key roles in viral assembly and uncoating. It also plays non-structural roles related to transcription modulation through nucleo-cytoplasmic shuttling and nucleic acid binding ability. We dissected the structural and conformational changes underlying the switch between multiple functionalities, identifying Ca 2+ binding as a key factor. To this end, we tackled the analysis of M's conformational stability and equilibria. While in silico calculations predict two potential calcium binding sites per protomer, purified RSV-M dimer contains only one strongly bound calcium ion per protomer. Incubation of RSV-M in the presence of excess Ca 2+ leads to an increase in the thermal stability, confirming additional Ca 2+ binding sites. Moreover, mild denaturant concentrations trigger the formation of higher order oligomers which are otherwise prevented under Ca 2+ saturation conditions, in line with the stabilizing effect of the additional low affinity binding site. On the other hand, Ca 2+ removal by chelation at pH 7.0 causes a substantial decrease in the thermal stability leading to the formation of amorphous, spherical-like aggregates, as assessed by TEM. Even though the Ca 2+ content modulates RSV-M oligomerization propensity, it does affect its weak RNA binding ability. RSV-M undergoes a substantial conformational change at pHs 4.0 to 5.0 that results in the exposure of hydrophobic surfaces, an increase beta sheet content but burial of tryptophan residues. While low ionic strength promotes dimer dissociation at pH 4.0, physiological concentrations of NaCl lead to the formation of soluble oligomers smaller than 400 kDa at pH 4.0 or insoluble aggregates with tubular morphology at pH 5.0, supporting a fine tuning by pH. Furthermore, the dissociation constants estimated for the low- and high affinity calcium binding sites are 13 μM and 58 nM, respectively, suggesting an intracellular calcium sensing mechanism of RSV-M upon infection. We uncover a finely tuned interplay between calcium binding, ionic strength, and pH changes compatible with the different cellular compartments where M plays key roles, revealing diverse conformational equilibria, oligomerization, and high order structures, required to stabilize the virion particle by a layer of molecules positioned between the membrane and the nucleocapsid.
(Copyright © 2022 Elsevier Inc. All rights reserved.)
Databáze: MEDLINE
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