Gene 4 of Marburg virus, strain Musoke, was subjected to nucleotide sequence analysis. It is 2,844 nucleotides long and extends from genome position 5821 to position 8665 (EMBL Data Library, emnew: MVREPCYC [accession no. Z12132]). The gene is flanked by transcriptional signal sequences (start signal, 3'-UACUUCUUGUAAUU-5'; termination signal, 3'-UAAUUCUUUUU-5') which are conserved in all Marburg virus genes. The major open reading frame encodes a polypeptide of 681 amino acids (M(r), 74,797). After in vitro transcription and translation, as well as expression in Escherichia coli, this protein was identified by its immunoreactivity with specific antisera as the unglycosylated form of the viral membrane glycoprotein (GP). The GP is characterized by the following four different domains: (i) a hydrophobic signal peptide at the amino terminus (1 to 18), (ii) a predominantly hydrophilic external domain (19 to 643), (iii) a hydrophobic transmembrane anchor (644 to 673), and (iv) a small hydrophilic cytoplasmic tail at the carboxy terminus (674 to 681). Amino acid analysis indicated that the signal peptide is removed from the mature GP. The GP therefore has the structural features of a type I transmembrane glycoprotein. The external domain of the protein has 19 N-glycosylation sites and several clusters of hydroxyamino acids and proline residues that are likely to be the attachment sites for about 30 O-glycosidic carbohydrate chains. The region extending from positions 585 to 610 shows significant homology to a domain observed in the envelope proteins of several retroviruses and Ebola virus that has been suspected to be responsible for immunosuppressive properties of these viruses. A second open reading frame of gene 4 has the coding capacity for an unidentified polypeptide 112 amino acids long.