High throughput screening platform for enzymes, pathways and strains using ‘Smart’ Microbial Cell Technology
While ‘Design’ and ‘Build’ steps of DBTL cycle for bio-based products show high throughput, the ‘Test’ step remains a bottleneck due to low efficiency of the current screening technologies. We have developed Smart Microbial Cell technology that utilizes a custom-design sensor-reporter system for a given small molecule of interest (value-added chemical, industrial precursor) in the host strain harboring variations in a single gene, or a pathway, or at the genome level. Based on the production efficiency of enzyme/pathway/strain (determined by genotype), there is a correlated response from the sensor-reporter system (phenotype), offering an easy screening technique using flow cytometry or by simply plating them on a solid phase and visualizing under an illuminator. This capability is unique as it provides the ability to incorporate computational protein design to engineer sensors or intelligent library design for evolution of enzymes. The existing capability enhances the efficiency of the ‘Test’ step of DBTL cycle for bio-based products. Product strains can be optimized in a high throughput manner. 102-106 variants can be screened in a single day.
References and Additional Information:
Jha RK, Narayanan N, Pandey N, Bingen J, Kern TL, Johnson CW, Strauss CEM, Beckham GT, Hennelly SP, Dale T. Sensor-enabled alleviation of product inhibition in chorismate pyruvate-lyase. ACS Synthetic Biology (2019), 10.1021/acssynbio.8b00465.
Jha RK, Bingen JM, Johnson CW, Kern TL, Khanna P, Trettel DS, Strauss CEM, Beckham GT, Dale T. A protocatechuate biosensor for Pseudomonas putida KT2440 via promoter and protein evolution. Manuscript accepted in Metabolic Engineering Communications (2018), 6, 33-38.
Los Alamos National Laboratory