Photonic activation of disulfide bridges achieves oriented protein immobilization on biosensor surfaces.
Photonic induced immobilization is a novel technology that results in spatially oriented and spatially localized covalent coupling of biomolecules onto thiol-reactive surfaces. Immobilization using this technology has been achieved for a wide selection of proteins, such as hydrolytic enzymes (lipases/esterases, lysozyme), proteases (human plasminogen), alkaline phosphatase, immunoglobulins' Fab fragment (e.g., antibody against PSA [prostate specific antigen]), Major Histocompability Complex class I protein, pepsin, and trypsin. The reaction mechanism behind the reported new technology involves "photonic activation of disulfide bridges," i.e., light-induced breakage of disulfide bridges in proteins upon UV illumination of nearby aromatic amino acids, resulting in the formation of free, reactive thiol groups that will form covalent bonds with thiol-reactive surfaces (see Fig. 1). Interestingly, the spatial proximity of aromatic residues and disulfide bridges in proteins has been preserved throughout molecular evolution. The new photonic-induced method for immobilization of proteins preserves the native structural and functional properties of the immobilized protein, avoiding the use of one or more chemical/thermal steps. This technology allows for the creation of spatially oriented as well as spatially defined multiprotein/DNA high-density sensor arrays with spot size of 1 microm or less, and has clear potential for biomedical, bioelectronic, nanotechnology, and therapeutic applications.
Neves-Petersen
Maria Teresa
MT
NanoBiotechnology Group, Department of Physics and Nanotechnology, Aalborg University, Aalborg, Denmark.
Snabe
Torben
T
Klitgaard
Søren
S
Duroux
Meg
M
Petersen
Steffen B
SB
D000911:Antibodies, Monoclonal / Q000737:chemistry; D015374:Biosensing Techniques; D004220:Disulfides / Q000737:chemistry*; D004800:Enzymes, Immobilized / Q000737:chemistry*; D005453:Fluorescence; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry; D008027:Light; D036103:Nanotechnology / Q000295:instrumentation* / Q000379:methods; D010777:Photochemistry; D040081:Protein Array Analysis; D011506:Proteins / Q000737:chemistry*; D011791:Quartz; D014466:Ultraviolet Rays
D000911:Antibodies, Monoclonal; D004220:Disulfides; D004800:Enzymes, Immobilized; D007140:Immunoglobulin Fab Fragments; D011506:Proteins; D011791:Quartz