Programmable assembly of a metabolic pathway enzyme in a pre-packaged reusable bioMEMS device.

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TitleProgrammable assembly of a metabolic pathway enzyme in a pre-packaged reusable bioMEMS device.
Publication TypeJournal Article
Year of Publication2008
AuthorsLuo, X, Lewandowski, AT, Yi, H, Payne, GF, Ghodssi, R, Bentley, WE, Rubloff, GW
JournalLab Chip
Volume8
Issue3
Pagination420-30
Date Published2008 Mar
ISSN1473-0197
KeywordsElectrodes, Enzyme Stability, Enzymes, Miniaturization, Optics and Photonics
Abstract

We report a biofunctionalization strategy for the assembly of catalytically active enzymes within a completely packaged bioMEMS device, through the programmed generation of electrical signals at spatially and temporally defined sites. The enzyme of a bacterial metabolic pathway, S-adenosylhomocysteine nucleosidase (Pfs), is genetically fused with a pentatyrosine "pro-tag" at its C-terminus. Signal responsive assembly is based on covalent conjugation of Pfs to the aminopolysaccharide, chitosan, upon biochemical activation of the pro-tag, followed by electrodeposition of the enzyme-chitosan conjugate onto readily addressable sites in microfluidic channels. Compared to traditional physical entrapment and surface immobilization approaches in microfluidic environments, our signal-guided electrochemical assembly is unique in that the enzymes are assembled under mild aqueous conditions with spatial and temporal programmability and orientational control. Significantly, the chitosan-mediated enzyme assembly can be reversed, making the bioMEMS reusable for repeated assembly and catalytic activity. Additionally, the assembled enzymes retain catalytic activity over multiple days, demonstrating enhanced enzyme stability. We envision that this assembly strategy can be applied to rebuild metabolic pathways in microfluidic environments for antimicrobial drug discovery.

DOI10.1039/b713756g
Alternate JournalLab Chip
PubMed ID18305860