Small P particles formed by the Taiwan-native norovirus P domain overexpressed in Komagataella pastoris
Journal
Applied Microbiology and Biotechnology
Journal Volume
102
Journal Issue
22
Pages
9707-9718
Date Issued
2018
Author(s)
Abstract
The protrusion (P) domain of the major structural protein VP1 of norovirus (NoV) is critical for the host’s immune response and receptor binding. Most heterologous P domains expressed in Escherichia coli or Komagataella pastoris (formally known as Pichia pastoris) form P particles consisting of 24 P monomers formed through intermolecular contact in the P regions and an end-linked cysteine tag. The small P particle is only found in P domains with terminal modifications. In this study, the NoV P domain of the most predominant NoV strain GII.4 isolated from Taiwan was expressed in K. pastoris. A high yield of NoV P was obtained using the high-cell density fermentation process in K. pastoris. A large amount of the small P particles and the trimer and dimer complexes formed by 12, 6, and 2 P monomers were observed in both the expression of the NoV P-His and P containing cysteine tag at the N-terminus. Dynamic light scattering and transmission electron microscopy analysis of the purified NoV P-His and P revealed that most of these small P particles are triangle-, square-, and ring-shaped with a diameter of 14–15?nm. The binding ability of purified NoV P-His and P to human histo-blood group antigen was confirmed by a saliva-binding assay. Without terminal modification, small P particles were formed in our study. The amino acid sequence analysis showed only four different amino acids (residue 84, 119, 136, and 313) between the P domain in this study and other investigated GII.4 strains suggesting that these amino acids might play an important role in the P particle formation. The small P particles formed by the Taiwan-native norovirus P domain overexpressed in K. pastoris may provide further information for morphogenesis studies and vaccine development. ? 2018, Springer-Verlag GmbH Germany, part of Springer Nature.
Subjects
Dimers; Escherichia coli; High resolution transmission electron microscopy; Light scattering; Light transmission; Monomers; Purification; Strain; Transmission electron microscopy; Yeast; Amino acid sequence analysis; Blood group antigen; High cell density fermentations; Intermolecular contacts; Norovirus; Pichia Pastoris; Terminal modifications; Transmission electron; Amino acids; cysteine; dimer; monomer; protein VP1; structural protein; viral protein; antigen; chemical binding; coliform bacterium; fermentation; gene expression; immune response; particulate matter; protein; virus; amino acid sequence; amino terminal sequence; Article; enzyme linked immunosorbent assay; fermentation; Komagataella pastoris; liquid chromatography-mass spectrometry; nonhuman; Norovirus; particle size; photon correlation spectroscopy; polyacrylamide gel electrophoresis; protein expression; saliva; transmission electron microscopy; virus particle; Western blotting; budding yeast; chemistry; gene expression; genetics; human; metabolism; Norovirus; protein domain; protein motif; Taiwan; Escherichia coli; Norovirus; Pichia pastoris; Amino Acid Motifs; Gene Expression; Humans; Norovirus; Protein Domains; Saccharomycetales; Taiwan; Viral Structural Proteins
SDGs
Other Subjects
Dimers; Escherichia coli; High resolution transmission electron microscopy; Light scattering; Light transmission; Monomers; Purification; Strain; Transmission electron microscopy; Yeast; Amino acid sequence analysis; Blood group antigen; High cell density fermentations; Intermolecular contacts; Norovirus; Pichia Pastoris; Terminal modifications; Transmission electron; Amino acids; cysteine; dimer; monomer; protein VP1; structural protein; viral protein; antigen; chemical binding; coliform bacterium; fermentation; gene expression; immune response; particulate matter; protein; virus; amino acid sequence; amino terminal sequence; Article; enzyme linked immunosorbent assay; fermentation; Komagataella pastoris; liquid chromatography-mass spectrometry; nonhuman; Norovirus; particle size; photon correlation spectroscopy; polyacrylamide gel electrophoresis; protein expression; saliva; transmission electron microscopy; virus particle; Western blotting; budding yeast; chemistry; gene expression; genetics; human; metabolism; Norovirus; protein domain; protein motif; Taiwan; Escherichia coli; Norovirus; Pichia pastoris; Amino Acid Motifs; Gene Expression; Humans; Norovirus; Protein Domains; Saccharomycetales; Taiwan; Viral Structural Proteins
Type
journal article