Engineering Dual-Input Glucose- and Temperature-Sensitive Lysis Circuits in Corynebacterium glutamicum for Efficient Intracellular Product Recovery

<i>Corynebacterium glutamicum</i> is a versatile microbial cell factory, but efficient recovery of intracellular macromolecules remains a major challenge. In this study, we engineered environmentally controllable lysis systems to enable programmable product release. A glucose-responsive module, combining the <i>cg3195</i> promoter with phage-derived holin-endolysin genes (<i>clg51-50</i>), triggered lysis when extracellular glucose dropped below 0.19-0.36 g/L. A separate temperature-inducible system employing the <i>cI857-CJ1OX2</i> module activated lysis at 42 °C. These modules were further integrated into a dual-input <i>AND-gate</i> circuit, enhancing regulatory precision and suppressing premature lysis, with additional operator copies allowing temporal tuning of induction. Functional validation using fluorescence, cell density measurements, and scanning electron microscopy confirmed robust, tunable responses under defined environmental cues. Collectively, these programmable lysis systems demonstrate that stimulus-responsive genetic circuits can be rationally designed to control cell disruption, providing a promising approach to streamline downstream processing and reduce extraction costs in industrial fermentation of <i>Corynebacterium glutamicum</i>.