tetano
Editor, Senior Moderator
Biochim Biophys Acta Gen Subj. 2019 Feb 27. pii: S0304-4165(19)30049-2. doi: 10.1016/j.bbagen.2019.02.015. [Epub ahead of print]
[h=1]Direct visualization of avian influenza H5N1 Hemagglutinin precursor and its conformational change by high-speed atomic force microscopy.[/h] Lim KS[SUP]1[/SUP], Mohamed MS[SUP]2[/SUP], Wang H[SUP]3[/SUP], Hartono[SUP]3[/SUP], Hazawa M[SUP]4[/SUP], Kobayashi A[SUP]1[/SUP], Voon DC[SUP]5[/SUP], Kodera N[SUP]1[/SUP], Ando T[SUP]1[/SUP], Wong RW[SUP]6[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] [h=4]BACKGROUND:[/h] Hemagglutinin (HA) of influenza A is one of the key virulence factors that mediates the release of viral components in host cells. HA is initially synthesized as a trimeric precursor (HA0) and then it is cleaved by proteases to become a functional HA. Low pH induces irreversible conformational changes in both HA0 and HA but only HA is fusion compatible. Here, we used high-speed atomic force microscopy (HS-AFM) to record conformational changes in HA0 trimers (H5N1) from neutral to acidic conditions at a millisecond scale.
[h=4]METHODS:[/h] Purified HA0 protein was diluted with either neutral Tris-HCl (pH: 7.4) or acetic acid-titrated Tris-HCl (pH 5.0) and then loaded onto bare mica. Neutral or acidic Tris-HCl was used as the scanning buffer. HS-AFM movies were recorded and processed using Image J software.
[h=4]RESULTS:[/h] The conformation of HA0[SUB]neutral[/SUB] visualized using HS-AFM was comparable to the HA trimer structures depicted in the PDB data and the AFM simulator. HA0 underwent rapid conformational changes under low pH condition. The circularity and area of HA0[SUB]acid[/SUB] were significantly higher than in HA0[SUB]neutral[/SUB]. In contrast, the height of HA0[SUB]acid[/SUB] was significantly lower than in HA0[SUB]neutral.[/SUB] CONCLUSIONS: We have captured real-time images of the native HA0 trimer structure under physiological conditions using HS-AFM. By analyzing the images, we confirm that HA0 trimer is sensitive to acidic conditions.
[h=4]GENERAL SIGNIFICANCE:[/h] The dynamic nature of the HA structure, particularly in the host endosome, is essential for H5N1 infectivity. Understanding this acidic behavior is imperative for designing therapeutic strategies against H5N1. This article reports a sophisticated new tool for studying the spatiotemporal dynamics of the HA precursor protein.
Copyright ? 2019. Published by Elsevier B.V.
[h=4]KEYWORDS:[/h] Conformational dynamic; HA0 trimer; Hemagglutinin; High-speed atomic force microscopy; Influenza A
PMID: 30825615 DOI: 10.1016/j.bbagen.2019.02.015
[h=1]Direct visualization of avian influenza H5N1 Hemagglutinin precursor and its conformational change by high-speed atomic force microscopy.[/h] Lim KS[SUP]1[/SUP], Mohamed MS[SUP]2[/SUP], Wang H[SUP]3[/SUP], Hartono[SUP]3[/SUP], Hazawa M[SUP]4[/SUP], Kobayashi A[SUP]1[/SUP], Voon DC[SUP]5[/SUP], Kodera N[SUP]1[/SUP], Ando T[SUP]1[/SUP], Wong RW[SUP]6[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] [h=4]BACKGROUND:[/h] Hemagglutinin (HA) of influenza A is one of the key virulence factors that mediates the release of viral components in host cells. HA is initially synthesized as a trimeric precursor (HA0) and then it is cleaved by proteases to become a functional HA. Low pH induces irreversible conformational changes in both HA0 and HA but only HA is fusion compatible. Here, we used high-speed atomic force microscopy (HS-AFM) to record conformational changes in HA0 trimers (H5N1) from neutral to acidic conditions at a millisecond scale.
[h=4]METHODS:[/h] Purified HA0 protein was diluted with either neutral Tris-HCl (pH: 7.4) or acetic acid-titrated Tris-HCl (pH 5.0) and then loaded onto bare mica. Neutral or acidic Tris-HCl was used as the scanning buffer. HS-AFM movies were recorded and processed using Image J software.
[h=4]RESULTS:[/h] The conformation of HA0[SUB]neutral[/SUB] visualized using HS-AFM was comparable to the HA trimer structures depicted in the PDB data and the AFM simulator. HA0 underwent rapid conformational changes under low pH condition. The circularity and area of HA0[SUB]acid[/SUB] were significantly higher than in HA0[SUB]neutral[/SUB]. In contrast, the height of HA0[SUB]acid[/SUB] was significantly lower than in HA0[SUB]neutral.[/SUB] CONCLUSIONS: We have captured real-time images of the native HA0 trimer structure under physiological conditions using HS-AFM. By analyzing the images, we confirm that HA0 trimer is sensitive to acidic conditions.
[h=4]GENERAL SIGNIFICANCE:[/h] The dynamic nature of the HA structure, particularly in the host endosome, is essential for H5N1 infectivity. Understanding this acidic behavior is imperative for designing therapeutic strategies against H5N1. This article reports a sophisticated new tool for studying the spatiotemporal dynamics of the HA precursor protein.
Copyright ? 2019. Published by Elsevier B.V.
[h=4]KEYWORDS:[/h] Conformational dynamic; HA0 trimer; Hemagglutinin; High-speed atomic force microscopy; Influenza A
PMID: 30825615 DOI: 10.1016/j.bbagen.2019.02.015