Characteristics of hCoV-NL63 open reading frame 3 and comparison to homologous genes in other coronaviruses. The sequence of ORF 3 (GenBank accession no. AY567487.2) was analyzed using BLAST and MEGA4. (A), localization of ORF 3 within the hCoV-NL63 genome and comparison of nucleotide (nt) identity based on multiple sequence alignments with prototype strains of CoV groups alpha, beta, gamma. Note that IBV ORF 3a and b were fused to one ORF 3ab. (B), Summarized results of in-silico analysis on membrane topology and glycosylation (refer to Materials and Methods section). Predicted N-linked glycosylation sites are indicated by an "N" at the respective localizations with an index number indentifying the amino acid position. No O linked glycosylation sites were predicted.