3 years ago

Self-Immobilizing Fusion Enzymes for Compartmentalized Biocatalysis

Self-Immobilizing Fusion Enzymes for Compartmentalized Biocatalysis
Teresa Burgahn, Christof M. Niemeyer, Sabrina Gallus, Marc Skoupi, Kersten S. Rabe, Theo Peschke, Ishtiaq Ahmed
The establishment of microfluidic enzyme cascades is a topical field of research and development, which is currently hampered by the lack of methodologies for mild and efficient immobilization of isolated enzymes. We here describe the use of self-immobilizing fusion enzymes for the modular configuration of microfluidic packed-bed reactors. Specifically, three different enzymes, the (R)-selective alcohol dehydrogenase LbADH, the (S)-selective methylglyoxal reductase Gre2p and the NADP(H) regeneration enzyme glucose 1-dehydrogenase GDH, were genetically fused with streptavidin binding peptide, Spy and Halo-based tags, to enable their specific and directional immobilization on magnetic microbeads coated with complementary receptors. The enzyme-modified beads were loaded in four-channel microfluidic chips to create compartments that have the capability for either (R)- or (S)-selective reduction of the prochiral CS-symmetrical substrate 5-nitrononane-2,8-dione (NDK). Analysis of the isomeric hydroxyketone and diol products by chiral HPLC was used to quantitatively characterize the performance of reactors configured with different amounts of the enzymes. Long operating times of up to 14 days indicated stable enzyme immobilization and the general robustness of the reactor. Even more important, by fine-tuning of compartment size and loading, the overall product distribution could be controlled to selectively produce a single meso diol with nearly quantitative conversion (>95%) and excellent stereoselectivity (d.r. > 99:1) in a continuous flow process. We believe that our concept will be expandable to a variety of other biocatalytic or chemo-enzymatic cascade reactions.

Publisher URL: http://dx.doi.org/10.1021/acscatal.7b02230

DOI: 10.1021/acscatal.7b02230

You might also like
Discover & Discuss Important Research

Keeping up-to-date with research can feel impossible, with papers being published faster than you'll ever be able to read them. That's where Researcher comes in: we're simplifying discovery and making important discussions happen. With over 19,000 sources, including peer-reviewed journals, preprints, blogs, universities, podcasts and Live events across 10 research areas, you'll never miss what's important to you. It's like social media, but better. Oh, and we should mention - it's free.

  • Download from Google Play
  • Download from App Store
  • Download from AppInChina

Researcher displays publicly available abstracts and doesn’t host any full article content. If the content is open access, we will direct clicks from the abstracts to the publisher website and display the PDF copy on our platform. Clicks to view the full text will be directed to the publisher website, where only users with subscriptions or access through their institution are able to view the full article.