Journal: PLoS ONE
Article Title: Non-human primate orthologues of TMPRSS2 cleave and activate the influenza virus hemagglutinin
Figure Lengend Snippet: TMPRSS2, TMPRSS4 and HAT are conserved between humans and non-human primates. (A) Amino acid sequence alignment of human (NP_005647.3), rhesus macaque (XP_014988331.1), cynomolgus macaque (XP_015302312.1) and common marmoset (XP_008984973.1) TMPRSS2. Protein alignment was performed by using Vector NTI AlignX. Colors indicate amino acid identity (yellow), conservation (blue) and similarity (green). The catalytic triad is boxed. (B) For analysis of protease expression, 293T cells were transfected with plasmids encoding TMPRSS2, TMPRSS4 or HAT of the indicated species and equipped with an N-terminal myc antigenic tag. Empty plasmid (pCAGGS) served as a negative control. Protease expression in cell lysates was detected via Western blotting with anti-myc antibody. Due to more prominent expression of TMPRSS2 relative to TMPRSS4 and HAT proteins, 10 μl of lysates from TMPRSS2 expressing cells and 20 μl of lysates from TMPRSS4 and HAT expressing cells were loaded for separation by SDS gel-electrophoresis. The expression of β-actin was determined as a loading control. Filled triangles indicate zymogen forms, while empty triangles highlight cleavage products resulting from autocatalytic activation. The results were confirmed in at least two separate experiments.
Article Snippet: The alignments of the amino acid sequences of the human and NHP TTSPs were constructed using AlignX Vector NTI (ThermoFisher).
Techniques: HAT Assay, Sequencing, Plasmid Preparation, Expressing, Transfection, Negative Control, Western Blot, SDS-Gel, Electrophoresis, Activation Assay