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Investigation of pathways for the low-pH conformational transition in influenza hemagglutinin.

ABSTRACT: Targeted molecular dynamics simulations were used to study the conformational transition of influenza hemagglutinin (HA) from the native conformation to putative fusogenic or postfusion conformations populated at low pH. Three pathways for this conformational change were considered. Complete dissociation of the globular domains of HA was observed in one pathway, whereas smaller rearrangements were observed in the other two. The fusion peptides became exposed and moved toward the target membrane, although occasional movement toward the viral membrane was also observed. The effective energy profiles along the paths show multiple barriers. The final low-pH structures, which are consistent with available experimental data, are comparable in effective energy to native HA. As a control, the uncleaved precursor HA0 was also forced along the same pathway. In this case both the final energy and the energy barrier were much higher than in the cleaved protein. This study suggests that 1) as proposed, the native conformation is the global minimum energy conformation for the uncleaved precursor but a metastable state for cleaved HA; 2) the spring-loaded conformational change is energetically plausible in full-length HA; and 3) complete globular domain dissociation is not necessary for extension of the coiled coil and fusion peptide exposure, but the model with complete dissociation has lower energy.

SUBMITTER: Madhusoodanan M

PROVIDER: S-EPMC1302762 | biostudies-literature | 2003 Mar

REPOSITORIES: biostudies-literature

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Investigation of pathways for the low-pH conformational transition in influenza hemagglutinin.

Madhusoodanan M M Lazaridis Themis T

Biophysical journal 20030301 3

Targeted molecular dynamics simulations were used to study the conformational transition of influenza hemagglutinin (HA) from the native conformation to putative fusogenic or postfusion conformations populated at low pH. Three pathways for this conformational change were considered. Complete dissociation of the globular domains of HA was observed in one pathway, whereas smaller rearrangements were observed in the other two. The fusion peptides became exposed and moved toward the target membrane, ...[more]

PMID: 12609895

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Project description:The M2 protein from influenza A plays important roles in its viral cycle. It contains a single transmembrane helix, which oligomerizes into a homotetrameric proton channel that conducts in the low-pH environment of the host-cell endosome and Golgi apparatus, leading to virion uncoating at an early stage of infection. We studied conformational rearrangements that occur in the M2 core transmembrane domain residing on the lipid bilayer, flanked by juxtamembrane residues (M2TMD21-49 fragment), upon its interaction with amantadine drug at pH 5.5 when M2 is conductive. We also tested the role of specific mutation and lipid chain length. Electron spin resonance (ESR) spectroscopy and electron microscopy were applied to M2TMD21-49, labeled at the residue L46C with either nitroxide spin-label or Nanogold® reagent, respectively. Electron microscopy confirmed that M2TMD21-49 reconstituted into DOPC/POPS at 1:10,000 peptide-to-lipid molar ratio (P/L) either with or without amantadine, is an admixture of monomers, dimers, and tetramers, confirming our model based on a dimer intermediate in the assembly of M2TMD21-49. As reported by double electron-electron resonance (DEER), in DOPC/POPS membranes amantadine shifts oligomer equilibrium to favor tetramers, as evidenced by an increase in DEER modulation depth for P/L's ranging from 1:18,000 to 1:160. Furthermore, amantadine binding shortens the inter-spin distances (for nitroxide labels) by 5-8 Å, indicating drug induced channel closure on the C-terminal side. No such effect was observed for the thinner membrane of DLPC/DLPS, emphasizing the role of bilayer thickness. The analysis of continuous wave (cw) ESR spectra of spin-labeled L46C residue provides additional support to a more compact helix bundle in amantadine-bound M2TMD 21-49 through increased motional ordering. In contrast to wild-type M2TMD21-49, the amantadine-bound form does not exhibit noticeable conformational changes in the case of G34A mutation found in certain drug-resistant influenza strains. Thus, the inhibited M2TMD21-49 channel is a stable tetramer with a closed C-terminal exit pore. This work is aimed at contributing to the development of structure-based anti-influenza pharmaceuticals.

Project description:The conformational change of the influenza virus hemagglutinin (HA) protein mediating the fusion between the virus envelope and the endosomal membrane was hypothesized to be induced by protonation of specific histidine residues since their pKas match the pHs of late endosomes (pK(a) of ? 6.0). However, such critical key histidine residues remain to be identified. We investigated the highly conserved His184 at the HA1-HA1 interface and His110 at the HA1-HA2 interface of highly pathogenic H5N1 HA as potential pH sensors. By replacing both histidines with different amino acids and analyzing the effect of these mutations on conformational change and fusion, we found that His184, but not His110, plays an essential role in the pH dependence of the conformational change of HA. Computational modeling of the protonated His184 revealed that His184 is central in a conserved interaction network possibly regulating the pH dependence of conformational change via its pKa. As the propensity of histidine to get protonated largely depends on its local environment, mutation of residues in the vicinity of histidine may affect its pK(a). The HA of highly pathogenic H5N1 viruses carries a Glu-to-Arg mutation at position 216 close to His184. By mutation of residue 216 in the highly pathogenic as well as the low pathogenic H5 HA, we observed a significant influence on the pH dependence of conformational change and fusion. These results are in support of a pK(a)-modulating effect of neighboring residues.The main pathogenic determinant of influenza viruses, the hemagglutinin (HA) protein, triggers a key step of the infection process: the fusion of the virus envelope with the endosomal membrane releasing the viral genome. Whereas essential aspects of the fusion-inducing mechanism of HA at low pH are well understood, the molecular trigger of the pH-dependent conformational change inducing fusion has been unclear. We provide evidence that His184 regulates the pH dependence of the HA conformational change via its pK(a). Mutations of neighboring residues which may affect the pK(a) of His184 could play an important role in virus adaptation to a specific host. We suggest that mutation of neighboring residue 216, which is present in all highly pathogenic phenotypes of H5N1 influenza virus strains, contributed to the adaptation of these viruses to the human host via its effect on the pKa of His184.

Dataset Information

Investigation of pathways for the low-pH conformational transition in influenza hemagglutinin.

Publications

Investigation of pathways for the low-pH conformational transition in influenza hemagglutinin.

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