Synthesis and characterization of a recombinant myohemerythrin protein encoded by a synthetic gene

• Published in: Gene Vol. 99; no. 2; pp. 151 - 156
• Main Authors: Alexander, Hannah, Alexander, Stephen, Heffron, Fred, Fieser, Terry M., Hay, Beverly N., Getzoff, Elizabeth D., Tainer, John A., Lerner, Richard A.
• Format: Journal Article
• Language: English
• Published: Lausanne Elsevier B.V 15.03.1991; Amsterdam Elsevier; New York, NY
• Subjects: Amino Acid Sequence; Animals; Base Sequence; Biological and medical sciences; Cloning, Molecular; critical residues; Epitopes - immunology; Escherichia coli - genetics; Fundamental and applied biological sciences. Psychology; gene construction; Genes, Synthetic - genetics; Genes. Genome; Hemerythrin - analogs & derivatives; Hemerythrin - biosynthesis; Hemerythrin - genetics; Hemerythrin - immunology; immune recognition; In Vitro Techniques; Molecular and cellular biology; Molecular genetics; Molecular Sequence Data; Polychaeta - immunology; Precipitin Tests; protein antigenicity; Recombinant DNA; Recombinant Proteins - biosynthesis; Recombinant Proteins - genetics; Recombinant Proteins - immunology; synthetic oligodeoxyribonucleotide; Themiste zostericola; synthetic oligodeoxyribonucleotide; protein antigenicity; nt; TNA; mGPl-2 phage; Recombinant DNA; bp; Ap; reMHr; kb; wMHr; oligo; wt; aa; PBS; RBS; SDS; immune recognition; gene construction; PAGE; Rif; PMSF; MHr; IPTG; critical residues; MCS; ELISA; Marine environment; Proteins; Antigenicity; Antigenic determinant; Structure activity relation; Nucleotide sequence; Helmintha; Nemathelminthia; Method; Invertebrata; Chemical synthesis; Nematoda
• ISSN: 0378-1119; 1879-0038
• DOI: 10.1016/0378-1119(91)90121-Q

The antigenic epitopes of the myohemerythrin (MHr) molecule have been studied extensively. The critical amino acid residues responsible for its immune recognition have been identified by using synthetic peptides and the technique ofepitope scanning. To assess the true relevance of these techniques for determining the molecular mechanism of antigenic recognition and immunogenicity, the results obtained with isolated peptides should be tested in the context of the folded protein. To this end, we have designed and constructed a synthetic MHr gene, in modular form, which will allow subsequent alterations of nucleotide sequences encoding epitopes of interest. We have produced the recombinant protein at high level, and have shown by several criteria that it possesses the chemical, physical and immunological properties of the native worm protein. Thus, we have developed a valuable system for detailed immunological studies of the structure and chemistry required for antibody binding to protein.