Constant-pH MD simulations of the protonation-triggered conformational switching in diphtheria toxin translocation domain.

Autor: Oliveira NFB; BioISI - Instituto de Biosistemas e Ciências Integrativas, Faculdade de Ciências, Universidade de Lisboa, Lisboa, Portugal., Ladokhin AS; Department of Biochemistry and Molecular Biology, University of Kansas School of Medicine, Kansas City, Kansas, USA. Electronic address: aladokhin@kumc.edu., Machuqueiro M; BioISI - Instituto de Biosistemas e Ciências Integrativas, Faculdade de Ciências, Universidade de Lisboa, Lisboa, Portugal. Electronic address: machuque@ciencias.ulisboa.pt.
Jazyk: angličtina
Zdroj: Biophysical journal [Biophys J] 2024 Dec 17; Vol. 123 (24), pp. 4266-4273. Date of Electronic Publication: 2024 Aug 30.
DOI: 10.1016/j.bpj.2024.08.023
Abstrakt: Protonation of key residues in the diphtheria toxin translocation (T)-domain triggered by endosomal acidification is critical for inducing a series of conformational transitions critical for the cellular entry of the toxin. Previous experiments revealed the importance of histidine residues in modulating pH-dependent transitions. They suggested the presence of a "safety latch" preventing premature refolding of the T-domain by a yet poorly understood mechanism. Here, we used constant-pH molecular dynamics simulations to systematically investigate the protonation sequence in the wild-type T-domain and the following mutants: H223Q, H257Q, E259Q, and H223Q/H257Q. Comparison of these computational results with previous experimental data on T-domain stability and activity with the H-to-Q replacements confirms the role of H223 (pK a  = 6.5) in delaying the protonation of the main trigger, H257 (pK a  = 2.2 in the WT and pK a  = 4.9 in H223Q). Our calculations also reveal a very low pK a for a neighboring acidic residue E259, which does not get protonated even during simulations at pH 3. This residue also contributes to the formation of the safety latch, with the pK a of H257 increasing from 2.2 to 5.1 upon E259Q replacement. In contrast, the latter replacement has virtually no effect on the protonation of the H223. Thus, we conclude that the interplay of the protonation in the H223/H257/E259 triad has evolved to prevent triggering the accidental refolding of the T-domain by a fluctuation in the protonation of the main trigger at neutral pH, before the incorporation of the toxin inside the endosome. Subsequent acidification of the endosome overcomes the safety latch and triggers conformational switching via repulsion of H223 + and H257 + . This protonation/conformation relationship corroborates experimental findings and offers a detailed stepwise molecular description of the transition mechanism, which can be instrumental in optimizing the potential applications of the T-domain for targeted delivery of therapies to tumors and other diseased acidic tissues.
Competing Interests: Declaration of interests The authors declare no competing interests.
(Copyright © 2024 Biophysical Society. Published by Elsevier Inc. All rights reserved.)
Databáze: MEDLINE
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