[P008] Poster

1:00 PM - 1:50 PM JST(4:00 AM - 4:50 AM UTC)

[P04-458] Scalable and Semi-automated DBTL Workflows for High-throughput Enzyme Engineering in Biofoundries

Georgii Emelianov^1,2, Dong-Uk Song^1,3, Dae-Hee Lee^1,2,3, Bong Hyun Sung^1,2,3, Haseong Kim^1,2,3, Seung-Goo Lee^1,2,3, ○Hyewon Lee^1,2 (1. Korea Research Institute of Bioscience and Biotechnology (KRIBB) (Korea), 2. University of Science and Technology (UST) (Korea), 3. Korea Advanced Institute of Science & Technology (KAIST) (Korea))

Keywords:

Biofoundry,DBTL Workflow,Isoprene synthase,Methylococcus capsulatus Bath

Biofoundries integrating automation and data-driven design are increasingly recognized as effective platforms for accelerating enzyme engineering. Here, we present a modular, automation-enabled workflow for enzyme optimization, demonstrated using isoprene synthase (IspS), a rate-limiting enzyme in isoprene biosynthesis. By integrating coevolution-informed mutation design with automated build processes, we conducted iterative rounds of site-directed mutagenesis followed by NGS-based validation and experimental screening, evaluating ~100 variants per cycle with scalability to larger libraries. This workflow enabled the identification of IspS variants with up to a 4.5-fold improvement in catalytic efficiency and enhanced thermostability. Incorporation of the optimized enzyme into the methanotroph Methylococcus capsulatus Bath improved methane-to-isoprene conversion, achieving a maximum titer of 319.6 mg/L. Overall, this study highlights the effectiveness of automation-integrated DBTL workflows for enzyme engineering and supports their broader application in C1-based biomanufacturing.