Method: pcDNA-PB1, pcDNA-PB1a (catalytically inactive; D445A/D446A), pcDNA-PB2, pcDNA-PB2 K627E, pcDNA-PA, pcDNA-NP and pPOLI-NA, which encodes an NA vRNA segment, are derived from influenza A/WSN/33 virus and have been described previously.
Replication of a Dog-Origin H6N1 Influenza Virus in Cell Culture and Mice.
Method: 293T cells were transfected with chANP32A-X1 (0.5 mug/well) using Lipofectamine 2000 (Invitrogen) for 24 h, infected with PR8-PB2 K627E virus for 1 h at 37 C (MOI = 0.01) and cultured for indicated time point.
Method: The PB2 K627E substitution of pBD-PB2 was performed by site-directed mutagenesis by PCR.
Method: The recombinant PR8 viruses carrying PB2 627
Discussion: Furthermore, we did not detect the chANP32A-X1 and huANP32A in the RNPs or cRNPs of PB2 K627E or 627K polymerase (Figure 8), suggesting that ANP32A transiently interacted with the trimeric polymerase before the generation of RNPs.
Identification of Key Amino Acids in the PB2 and M1 Proteins of H7N9 Influenza Virus That Affect Its Transmission in Guinea Pigs.
Abstract: We found that the neuraminidase (NA) of the nontransmissible virus A/chicken/Shanghai/S1053/2013 had low enzymatic activity that impaired the transmission of AH/1 virus, and three amino acid mutations-V292I and K627E in PB2 and D156E in M1-independently abolished the transmission of the AH/1 virus.
Result: Our transmission study indicated that the mutations V292I and K627E of PB2 inde
Result: and C); K627E in PB2 also significantly reduced the polymerase activity of the RNP complex of the AH/1 virus in 293T cells.
Human Clade 2.3.4.4 A/H5N6 Influenza Virus Lacks Mammalian Adaptation Markers and Does Not Transmit via the Airborne Route between Ferrets.
Result: In order to understand whether the high polymerase activity of A/H5N6 GZ/14 was the result of the presence of PB2-627K, the PB2 gene of A/H5N6 GZ/14 was mutated to the avian genotype (K627E).
The PB2-K627E mutation attenuates H3N2 swine influenza virus in cultured cells and in mice.
PMID: 29175013
2018
Research in veterinary science
Abstract: For the first time, our results showed that PB2-K627E mutation attenuates H3N2 swine influenza virus in mammalian cells and in mice, suggesting that PB2-627K is required for viral replication and pathogenicity of H3N2 swine influenza virus.
Abstract: To explore the potential role of PB2-K627E substitution in H3N2 swine influenza virus, the growth properties and pathogenicity between H3N2 swine influenza virus and its PB2-K627E mutant were compared.
Prerequisites for the acquisition of mammalian pathogenicity by influenza A virus with a prototypic avian PB2 gene.
Result: Furthermore, rPB2(PR8)-IIIE and rPB2(PR8)-IIIEA replicated less efficiently than rPR8 and rPB2(PR8)-K627E.
Result: Furthermore, the combination of III with 627E or 627E and 674A [PB2(PR8)-IIIE and PB2(PR8)-IIIEA] significantly decreased the polymerase activity compared to PB2(PR8)-III, PB2(PR8)-IIIA, and PB2(PR8)-K627E (P < 0.05).
Result: Moreover, the pathogenicity of rPB2(PR8)-IIIE was markedly attenuated; this variant caused no body weight loss and produced much lower virus titres in the lungs of infected mice than rPB2(PR8)-K627E and rPB2(PR8)-III (Table 3).
Result: Next, we assessed the pathogenicity of rPB2(PR8)-IIIE, rPB2(PR8)-III, and rPB2(PR8)-K
Identification of polymerase gene mutations that affect viral replication in H5N1 influenza viruses isolated from pigeons.
PMID: 27926816
2017
The Journal of general virology
Abstract: In contrast, the PB2-K627E and PA-K158R mutations had moderate effects: PB2-K627E decreased and PA-K158R increased polymerase activity.
PB2-588 V promotes the mammalian adaptation of H10N8, H7N9 and H9N2 avian influenza viruses.
Result: By contrast, the PB2 substitution V588A markedly decreased virus replication of JX436 although it was not significant when compared to the substitution K627E and V588A/K627E in both MDCK and HEK293T cells.
Result: By contrast, the single substitution V588A, similar to K627E in PB2 for the human-origin JX346, had a slight effect on the MLD50 value, virulence, and virus replication in lungs of infected mice (Table 3), but the substitution V588A coupled with K627E in PB2 attenuated the parental JX346 virus, evidenced by a significantly enhanced MLD50 value and no case fatality rate in infected mice
Novel Polymerase Gene Mutations for Human Adaptation in Clinical Isolates of Avian H5N1 Influenza Viruses.
Result: As shown in S6 Fig, PB2-K627E produced less vRNA than EG/D1 (wt), indicating that PB2-E627K increased the replication activity of EG/D1 (wt) polymerase.
Result: In this study, 293T cells were co-transfected with plasmids expressing EG/D1 PB2, PB1, PA or NP, each carrying the indicated mutations that had a significant effect on progeny vRNA production in human airway cells, including PB2-E627K, with the PB2-K627E back mutation, and a plasmid expressing cRNA as a template for replication (cRNA to vRNA synthesis).
IV mutation literature information.
PMID: 
2020
Nature
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