The cytoplasmic tail of influenza A/H1N1 virus hemagglutinin is β-structural

Khrustalev, Vladislav Victorovich; Kordyukova, Larisa V.; Arutyunyan, Alexander M.; Poboinev, Victor Vitoldovich; Khrustaleva, Tatyana Aleksandrovna; Stojarov, Aleksander Nicolaevich; Baratova, L. A.; Sapon, Alena S; Лугин, В. Г.

doi:10.1080/07391102.2020.1860827

Cited by 8 publications

(27 citation statements)

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“…This shows the additional impact of the HA anchoring segments on the formation of thick lipid-protein nanodomains. Recently, based on the results of studying the secondary structure of a series of synthetic peptides corresponding to the HA CT sequence (H1 subtype), we proposed a model of this region containing an antiparallel β-structure with a turnaround and invariant glycine residue [ 49 ]. The height/depth of the β-hairpin is ~ 1 nm, which is consistent with the widening of the lipid bilayer of proteoliposomes containing such HA peptide configuration for about 1 nm at each side after loading them with M1.…”

Section: Resultsmentioning

confidence: 99%

The Cytoplasmic Tail of Influenza A Virus Hemagglutinin and Membrane Lipid Composition Change the Mode of M1 Protein Association with the Lipid Bilayer

et al. 2021

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Influenza A virus envelope contains lipid molecules of the host cell and three integral viral proteins: major hemagglutinin, neuraminidase, and minor M2 protein. Membrane-associated M1 matrix protein is thought to interact with the lipid bilayer and cytoplasmic domains of integral viral proteins to form infectious virus progeny. We used small-angle X-ray scattering (SAXS) and complementary techniques to analyze the interactions of different components of the viral envelope with M1 matrix protein. Small unilamellar liposomes composed of various mixtures of synthetic or “native” lipids extracted from Influenza A/Puerto Rico/8/34 (H1N1) virions as well as proteoliposomes built from the viral lipids and anchored peptides of integral viral proteins (mainly, hemagglutinin) were incubated with isolated M1 and measured using SAXS. The results imply that M1 interaction with phosphatidylserine leads to condensation of the lipid in the protein-contacting monolayer, thus resulting in formation of lipid tubules. This effect vanishes in the presence of the liquid-ordered (raft-forming) constituents (sphingomyelin and cholesterol) regardless of their proportion in the lipid bilayer. We also detected a specific role of the hemagglutinin anchoring peptides in ordering of viral lipid membrane into the raft-like one. These peptides stimulate the oligomerization of M1 on the membrane to form a viral scaffold for subsequent budding of the virion from the plasma membrane of the infected cell.

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Section: Resultsmentioning

confidence: 99%

The Cytoplasmic Tail of Influenza A Virus Hemagglutinin and Membrane Lipid Composition Change the Mode of M1 Protein Association with the Lipid Bilayer

et al. 2021

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“…Interestingly, the thermal shift assay method was also evaluated as a tool to assess stability, but no significant shifts in the structure were observed at the conditions evaluated (data not shown). Using CD, it is also possible to evaluate other temperature ranges, pHs, the effect of cryopreservants, and perform batch-to-batch comparison (Kissmann et al, 2011;Khrustalev et al, 2020).…”

Section: Vlps Structure and Thermal Stabilitymentioning

confidence: 99%

Bioanalytics for Influenza Virus-Like Particle Characterization and Process Monitoring

Carvalho

Silva

Sousa

et al. 2022

Front. Bioeng. Biotechnol.

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Virus-like particles (VLPs) are excellent platforms for the development of influenza vaccine candidates. Nonetheless, their characterization is challenging due to VLPs’ unique biophysical and biochemical properties. To cope with such complexity, multiple analytical techniques have been developed to date (e.g., single-particle analysis, thermal stability, or quantification assays), most of which are rarely used or have been successfully demonstrated for being applicable for virus particle characterization. In this study, several biophysical and biochemical methods have been evaluated for thorough characterization of monovalent and pentavalent influenza VLPs from diverse groups (A and B) and subtypes (H1 and H3) produced in insect cells using the baculovirus expression vector system (IC-BEVS). Particle size distribution and purity profiles were monitored during the purification process using two complementary technologies — nanoparticle tracking analysis (NTA) and tunable resistive pulse sensing (TRPS). VLP surface charge at the selected process pH was also assessed by this last technique. The morphology of the VLP (size, shape, and presence of hemagglutinin spikes) was evaluated using transmission electron microscopy. Circular dichroism was used to assess VLPs’ thermal stability. Total protein, DNA, and baculovirus content were also assessed. All VLPs analyzed exhibited similar size ranges (90–115 nm for NTA and 129–141 nm for TRPS), surface charges (average of −20.4 mV), and morphology (pleomorphic particles resembling influenza virus) exhibiting the presence of HA molecules (spikes) uniformly displayed on M1 protein scaffold. Our data shows that HA titers and purification efficiency in terms of impurity removal and thermal stability were observed to be particle dependent. This study shows robustness and generic applicability of the tools and methods evaluated, independent of VLP valency and group/subtype. Thus, they are most valuable to assist process development and enhance product characterization.

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“…Previously we have offered a 3D model of the HA CT region for Influenza A/H1N1 virus in the form of a peptide [ 2 ]. The next exciting step of the study is the modeling of the HA CT interactions with matrix protein M1 at physiological pH of 7.4 in the cellular cytoplasm and at acidic pH of 5.0 in the lysosomes.…”

Section: Introductionmentioning

confidence: 99%

“…It is almost impossible to model in silico such a complex structure of a lipopeptide with stearic and palmitic acid residues incorporated into the lipid bilayer [ 14 ]. That is why previously we have decided to use synthetic peptides corresponding to the last C-terminal 14 or 15 residues of Influenza A/H1N1 virus hemagglutinin to study their secondary structure in solution using circular dichroism (CD) and multi-bounce Horizontal Attenuated Total Reflectance Fourier Transformed Infrared (HATR-FTIR) spectroscopy analysis [ 2 ]. To prevent the possibility of disulfide bonds formation we substituted all three cysteine residues in two of those peptides by acetaminomethylcysteine residues [ 2 ].…”

Section: Introductionmentioning

confidence: 99%

“…Four peptides (WI14, FI15, WI14-ACM, and FI15-ACM) have been synthesized. The CD spectroscopy analysis of their saturated solutions at pH 7.4 revealed the formation of an antiparallel beta structure in all studied peptides [ 2 ]. At the same time, according to the results of fluorescence spectra analysis, the peptides are capable to form oligomers [ 2 ].…”

Section: Introductionmentioning

confidence: 99%

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Peptide Models of the Cytoplasmic Tail of Influenza A/H1N1 Virus Hemagglutinin Expand Understanding its pH-Dependent Modes of Interaction with Matrix Protein M1

et al. 2023

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Influenza A virus hemagglutinin (HA) is a major virus antigen. No cryo-electron microscopy or X-ray data can be obtained for the HA intraviral (cytoplasmic) domain (CT) post-translationally modified with long fatty acid residues bound to three highly conserved cysteines. We recently proposed a model of HA CT of Influenza A/H1N1 virus possessing an antiparallel beta structure based on the experimental secondary structure analysis of four 14–15 amino acid long synthetic peptides, corresponding to the HA CT sequence, with free or acetaminomethylated cysteines. To dispel doubts about possible non-specific “amyloid-like” aggregation of those synthetic peptides in phosphate buffer solution, we have determined the order of oligomers based on blue native gel electrophoresis, membrane filtration, fluorescence spectroscopy and molecular modeling approaches. We have found that unmodified peptides form only low molecular weight oligomers, while modified peptides form both oligomers of low order similar to those found for unmodified peptides and high order conglomerates, which however are not of beta-amyloid-like fold. This study confirms that the beta structure previously detected by circular dichroism spectroscopy analysis is more likely the result of intrinsic propensity of the HA CT amino acid sequence than the consequence of aggregation. The structures of low order oligomers of the synthetic peptides were used for in silico experiments on modeling of HA CT interactions with matrix protein M1 at physiological and acidic pH levels and revealed two different areas of binding. Finally, tripeptides capable of blocking interactions between HA CT and M1 were proposed. Supplementary Information The online version of this articlecontains supplementary material available 10.1007/s10930-023-10101-z.

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The cytoplasmic tail of influenza A/H1N1 virus hemagglutinin is β-structural

Cited by 8 publications

References 75 publications

The Cytoplasmic Tail of Influenza A Virus Hemagglutinin and Membrane Lipid Composition Change the Mode of M1 Protein Association with the Lipid Bilayer

The Cytoplasmic Tail of Influenza A Virus Hemagglutinin and Membrane Lipid Composition Change the Mode of M1 Protein Association with the Lipid Bilayer

Bioanalytics for Influenza Virus-Like Particle Characterization and Process Monitoring

Peptide Models of the Cytoplasmic Tail of Influenza A/H1N1 Virus Hemagglutinin Expand Understanding its pH-Dependent Modes of Interaction with Matrix Protein M1

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