N-glycosylation of hCoV-NL63 ORF 3 protein. HCoV-NL63 ORF 3 protein with and without a C-terminal V5 tag, and with an N16Q exchange in the tagged version was in-vitro translated in presence of 35S-methionine. SARS-CoV M protein without a tag was translated in the same system as a control. Proteins were digested with endoglycosidase (Endo H) as indicated below each lane, subjected to SDS-PAGE, and visualized. Note the removal of the bands of increased molecular weight for the control and ORF 3 proteins, but not for the ORF 3 protein with an amino acid exchange at the hypothetical N-glycosylation site. Note also that extent of size reduction for the SARS-CoV M protein, which is known to have one N-terminal N-glycosylation site, is the same for the NL63 ORF 3 protein.