Molecular mechanisms for antibody-mediated modulation of peptide-displaying enzyme sensors.
The generation of molecular sensors based on peptide-displaying enzymes for the detection of antibodies or antigens represents an innovative field of protein engineering. The knowledge of the underlying molecular mechanisms of enzymatic modulation in such sensors would be of great importance for the rational construction and improvement of responsiveness of new peptide-enzyme molecules. Here we analyze the enzymatic characteristics of three different kinds of sensors based in engineered beta-galactosidase, alkaline phosphatase and beta-lactamase, to explore a common activation basis. We describe two different categories of enzyme sensors. In one of them, including only some modified beta-lactamases, the enzymatic activity is inhibited upon ligand binding and it seems to be caused by the steric coverage of the active site by the bound antibody. In a second group, embracing members of the three studied enzymes, the ability to be modulated upon effector binding depends on the ratio between the k(cat) of the engineered enzyme and the k(cat) of the intact enzyme. This proves a common mechanism for enzymatic modulation of enzyme biosensors that is probably caused by conformational effects induced by the bound antibody on the enzyme.
Ferrer-Miralles
N
N
Departament de Genètica i Microbiologia, Institut de Biologia Fonamental, Barcelona, 08193, Spain. nferrer@einstein.uab.es
Feliu
J X
JX
Villaverde
A
A
D000469:Alkaline Phosphatase / Q000737:chemistry / Q000378:metabolism*; D000906:Antibodies / Q000737:chemistry*; D015374:Biosensing Techniques; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D001616:beta-Galactosidase / Q000737:chemistry / Q000378:metabolism*; D001618:beta-Lactamases / Q000737:chemistry / Q000378:metabolism*
D000906:Antibodies; D010455:Peptides; D011994:Recombinant Proteins; D000469:Alkaline Phosphatase; D001616:beta-Galactosidase; D001618:beta-Lactamases