Synthesis in Escherichia coli of human adenovirus type 12 transforming proteins encoded by early region 1A 13S mRNA and 12S mRNA.
Human adenovirus (Ad)-encoded early region 1A (E1A) tumor (T) antigens have been implicated in the positive regulation of viral early genes, the positive and negative regulation of some cellular genes, and cell immortalization and transformation. To further study the Ad E1A T antigens and to facilitate their purification, we have cloned cDNA copies of the Ad12 E1A 13S mRNA and 12S mRNA downstream of a hybrid Escherichia coli trp-lac (tac) promoter. Up to 8% of the protein synthesized in E. coli cells transformed by each of the two different Ad12 E1A cDNA constructs were immunoprecipitated as a Mr 47,000 protein by antibody to a synthetic peptide encoded in the Ad12 E1A DNA sequence. Both proteins produced in E. coli appear to be authentic and complete Ad12 E1A T antigens because they possess (i) the Ad12 E1A NH2-terminal amino acid sequence predicted from the DNA sequence; (ii) the Ad12 E1A COOH-terminal sequence, as shown by immunoprecipitation with anti-peptide antibody; and (iii) a molecular weight and an acidic isoelectric point similar to that of the E1A T antigens synthesized in Ad12-infected and transformed mammalian cells. The T antigens were purified to near homogeneity in yields of 100-200 micrograms per g wet weight of transformed E. coli cells.
D000260:Adenoviruses, Human / Q000235:genetics* / Q000276:immunology; D000957:Antigens, Viral, Tumor / Q000235:genetics / Q000302:isolation & purification; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005814:Genes, Viral; D007526:Isoelectric Point; D008970:Molecular Weight; D012333:RNA, Messenger / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics
D000957:Antigens, Viral, Tumor; D012333:RNA, Messenger; D012367:RNA, Viral