title: Site-Selective Oxidative Coupling Reactions for the Attachment of Enzymes to Glass Surfaces through DNA-Directed Immobilization.
abstract: Enzymes are able to maintain remarkably high selectivity toward their substrates while still retaining high catalytic rates. By immobilizing enzymes onto surfaces we can heterogenize these biological catalysts, making it practical to study, use, and combine them in an easily controlled system. In this work, we developed a platform that allows for the simple and oriented immobilization of proteins through DNA-directed immobilization. First, we modified a glass surface with single-stranded DNA. We then site-selectively attached the complementary DNA strand to the N-terminus of a protein. Both DNA modifications were carried out using an oxidative coupling strategy, and the DNA strands served as easily tunable and reversible chemical handles to hybridize the protein-DNA conjugates onto the surface. We have used the aldolase enzyme as a model protein to conduct our studies. We characterized each step of the protein immobilization process using fluorescent reporters as well as atomic force microscopy. We also conducted activity assays on the surfaces with DNA-linked aldolase to validate that, despite being modified with DNA and undergoing subsequent immobilization, the enzyme was still able to retain its catalytic activity and the surfaces were reusable in subsequent cycles.
authors.affiliation: Department of Chemistry, University of California , Berkeley, California 94720-1460, United States.
authors.forename: Kanwal S
authors.initials: KS
authors.lastname: Palla
authors.affiliation: Department of Chemistry, University of California , Berkeley, California 94720-1460, United States.|Chemical Sciences Division, Lawrence Berkeley National Laboratories , Berkeley, California 94720-1460, United States , and.
authors.forename: Tyler J
authors.initials: TJ
authors.lastname: Hurlburt
authors.affiliation: Department of Chemistry, University of California , Berkeley, California 94720-1460, United States.|Materials Sciences Division, Lawrence Berkeley National Laboratories , Berkeley, California 94720-1460, United States.
authors.forename: Alexander M
authors.initials: AM
authors.lastname: Buyanin
authors.affiliation: Department of Chemistry, University of California , Berkeley, California 94720-1460, United States.|Chemical Sciences Division, Lawrence Berkeley National Laboratories , Berkeley, California 94720-1460, United States , and.|Materials Sciences Division, Lawrence Berkeley National Laboratories , Berkeley, California 94720-1460, United States.
authors.forename: Gabor A
authors.initials: GA
authors.lastname: Somorjai
authors.affiliation: Department of Chemistry, University of California , Berkeley, California 94720-1460, United States.|Materials Sciences Division, Lawrence Berkeley National Laboratories , Berkeley, California 94720-1460, United States.
authors.forename: Matthew B
authors.identifier: 0000-0003-2837-2538
authors.initials: MB
authors.lastname: Francis
mesh_terms: D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000378:metabolism*; D005634:Fructose-Bisphosphate Aldolase / Q000737:chemistry / Q000378:metabolism*; D005898:Glass / Q000737:chemistry; D061925:Immobilized Nucleic Acids / Q000737:chemistry / Q000378:metabolism*; D018625:Microscopy, Atomic Force; D008958:Models, Molecular; D015394:Molecular Structure; D010084:Oxidation-Reduction; D013499:Surface Properties
chemical_list: D004800:Enzymes, Immobilized; D061925:Immobilized Nucleic Acids; D004247:DNA; D005634:Fructose-Bisphosphate Aldolase