Functional analysis of amino acids at stalk/head interface of human parainfluenza virus type 3 hemagglutinin-neuraminidase protein in the membrane fusion process
Human parainfluenza virus type 3 (hPIV3) is an important respiratory pathogen that causes the majority of viral pneumonia of infants and young children. hPIV3 can infect host cells through the synergistic action of hemagglutinin-neuraminidase (HN) protein and the homotypic fusion (F) protein on the viral surface. HN protein plays a variety of roles during the virus invasion process, such as promoting viral particles to bind to receptors, cleaving sialic acid, and activating the F protein. Crystal structure research shows that HN tetramer adopted a “heads-down” conformation, at least two heads dimmer on flank of the four-helix bundle stalk, which forms a symmetrical interaction interface. The stalk region determines interactions and activation of F protein in specificity, and the heads in down position statically shield these residues. In order to make further research on the function of these amino acids at the hPIV3 HN stalk/head interface, fifteen mutations (8 sites from stalk and 7 sites from head) were engineered into this interface by site-directed mutagenesis in this study. Alanine substitution in this region of hPIV3 HN had various effects on cell fusion promotion, receptor binding, and neuraminidase activity. Besides, L151A also affected surface protein expression efficiency. Moreover, I112A, D120A, and R122A mutations of the stalk region that were masked by global head in down position had influence on the interaction between F and HN proteins.
KeywordsHuman parainfluenza virus type 3 Hemagglutinin-neuraminidase protein Stalk/head interface Mutations Membrane fusion
We gratefully acknowledge Dr. Ronald Iorio for providing the recombinant plasmid vectors and Dr. Bernard Moss for vTF7-3.
JJ performed all parts of experiments and wrote the manuscript. MC and YL helped in experimental design, JL and ZC helped in the analysis of the results. HW, LZ, YS, NL, and LC supervised this study. ZW directed the experiments and revised the manuscript.
This work was supported by a Grant from National Natural Science Foundation of China (No. 81672011) and the Fundamental Research Funds of Shandong University (2015JC044).
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Conflicts of interest
All authors in this paper declare they have no conflict of interest.
Informed consent was obtained from all individual participants included in the study.
Research involving human participants and/or animals
Present paper does not contain any studies with human participants or animals performed by any of the authors.