Online Raman spectroscopy-guided strategy for efficient production of DNA recombinase UvsX in Escherichia coli

The rapid development of recombinase polymerase amplification (RPA) has driven a growing demand for UvsX, the key DNA recombinase for this technique. During bioreactor cultivation for UvsX production in Escherichia coli, biomass and substrate are critical factors affecting its efficient expression. However, the real-time monitoring techniques for these parameters remain immature, leading to delayed process control, hindering stable high-level production and precise process optimization. Using an online Raman spectrometer, this study developed synchronous quantitative prediction models for critical process parameters by partial least squares (PLS). Robust multivariate regression models showed strong correlations between predicted and measured values for biomass (R² = 0.994), glucose (R² = 0.979), UvsX (R² = 0.989), and total product (R² = 0.983). Based on these predictive models, the study implemented online monitoring and feedback control of critical fermentation parameters, successfully raising UvsX and total product titer to 8.23 g/L and 9.17 g/L, representing increases of 33.17% and 27.54% compared with conventional strategies. This study achieved a marked improvement in UvsX titer and production efficiency by applying Raman spectroscopy as a simple and rapid tool for process monitoring and optimization, providing dependable technical support for scaling up and quality control of complex recombinant proteins.