Porcine submaxillary gland apomucin contains tandemly repeated, identical sequences of 81 residues

• Published in: The Journal of biological chemistry Vol. 263; no. 2; pp. 1081 - 1088
• Main Authors: Timpte, C S, Eckhardt, A E, Abernethy, J L, Hill, R L
• Format: Journal Article
• Language: English
• Published: Bethesda, MD Elsevier Inc 15.01.1988; American Society for Biochemistry and Molecular Biology
• Subjects: Amino Acid Sequence; Animals; Base Sequence; Biological and medical sciences; Biotechnology; Deoxyribonucleases, Type II Site-Specific; DNA - analysis; DNA Restriction Enzymes - metabolism; Fundamental and applied biological sciences. Psychology; Gastric Mucins; Genes. Genome; Genetic engineering; Genetic technics; Methods. Procedures. Technologies; Molecular and cellular biology; Molecular cloning; Molecular genetics; Molecular Sequence Data; Peptide Mapping; Peptides - analysis; Peptides - genetics; Recombinant Fusion Proteins - analysis; Repetitive Sequences, Nucleic Acid; RNA, Messenger - analysis; Submandibular Gland - analysis; Swine; Nucleotide sequence; Apomucin; Submandibular gland; Glycoproteins; Secondary structure; Tandemly repeated sequence; Pig; Vertebrata; Mammalia; Mucin; Messenger RNA; Immunoblotting assay; Artiodactyla; Molecular cloning; Ungulata; Aminoacid sequence
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(19)35463-8

A lambda gt11 cDNA library, prepared from porcine submaxillary gland mRNA, was screened with anti-apomucin IgG, and five antibody-reactive phage were isolated. The phage with the largest cDNA insert, designated lambda PSM103, was further characterized. Its fusion protein reacted with anti-apomucin IgG and was used to affinity purify antibodies that specifically reacted with apomucin, indicating that the protein shares antigenic determinants with apomucin. The nucleotide sequence of 1510 bases in the 3.7-kilobase cDNA insert of lambda PSM103 has been established, thereby giving a deduced amino acid sequence of 503 residues in apomucin, or about 45% of the molecule. The deduced sequence of the apomucin polypeptide was found to contain 4.8 tandemly repeated, identical sequences of 81 residues each. The presence of these uniquely repeated sequences was confirmed by restriction endonuclease digestion of DNA derived from lambda PSM103. The repeat sequence was also confirmed in apomucin by the isolation of an 81-residue tryptic peptide with an amino acid composition and an amino-terminal amino acid sequence (up to 44 residues) identical to those of the tandem repeat. Moreover, the peptide was isolated in 760% yield, indicating that the tandem repeat occurs at least eight times in apomucin. The presence of such a long repetitive region in the gene for apomucin raises the possibility for considerable polymorphism in the gene and a corresponding size heterogeneity of apomucin. The predicted secondary structure of the 503 residues confirms the earlier proposal that apomucin is an extended, nonglobular polypeptide. Although the sequences around 192 serine and threonine residues have been established in apomucin, a recognition sequence for the N-acetylgalactosaminyltransferase that initiates glycosylation of apomucin is not evident, except that the glycosylated residues occur in turns.