Modulation of enzyme activity by antibody binding to an alkaline phosphatase-epitope hybrid protein.
An epitope from the HIV-1 gp120 protein V3 loop has been inserted onto the surface of bacterial alkaline phosphatase at different positions in the vicinity of the enzyme active site, creating hybrid proteins that can bind to an anti-gp120 monoclonal antibody. One of the hybrid proteins, API1, has a 13 amino acid V3 loop sequence inserted between residues 407 and 408 of alkaline phosphatase. The enzymatic activity of this protein is modulated upon antibody binding. API1 maintains the full activity of the wild type alkaline phosphatase but in the presence of the anti-gp120 antibody, the enzyme activity is inhibited by 40-50%. Thus, the hybrid enzyme can be used to detect the presence of antibody in solution. The concept of signalling proteins may have a wide application. Two models for the mechanism of modulation, steric hindrance and allosteric regulation, are discussed.
Brennan
C
C
Abbott Laboratories, IL 60064-3500.
Christianson
K
K
Surowy
T
T
Mandecki
W
W
D000469:Alkaline Phosphatase / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology; D001667:Binding, Competitive; D000939:Epitopes / Q000235:genetics / Q000276:immunology; D005818:Genetic Engineering; D015483:HIV Antibodies / Q000276:immunology*; D015699:HIV Envelope Protein gp120 / Q000235:genetics / Q000276:immunology*; D015497:HIV-1 / Q000235:genetics / Q000276:immunology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000939:Epitopes; D015483:HIV Antibodies; D015699:HIV Envelope Protein gp120; C088226:HIV envelope protein gp120 (302-324); D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D000469:Alkaline Phosphatase