Assembly and Functional Coordination of Two Families of Metabolic Organelles in Salmonella

Bacterial microcompartments (BMCs) are protein-based organelles that spatially organise metabolic pathways in prokaryotes, playing critical roles in enhancing metabolic processes and microbe fitness. Notably, many bacterial species possess multiple types of BMCs. While recent studies have advanced our knowledge about the assembly and function of individual BMC types, the mechanisms governing the coexistence and interplay of distinct BMC families within a single bacterial cell remain poorly understood. Here, we engineered Salmonella enterica serovar Typhimurium LT2 to co-express native 1,2-propanediol utilisation (Pdu) BMCs and synthetic α-carboxysomes (α-CBs), providing a unique platform for dissecting their assembly dynamics and functional crosstalk. By exploiting super-resolution fluorescence imaging, electron microscopy, biochemical and enzymatic assays, our studies demonstrate the formation of hybrid BMCs through the exchange of shell proteins between Pdu BMCs and α-CBs, whereas cargo proteins exhibit only limited compatibility, highlighting the specificity of encapsulation mechanisms. Furthermore, the generated hybrid BMCs display altered mobility and enzymatic activities, revealing emergent properties arising from shell protein interchangeability. Our findings provide insights into the inherent structural plasticity and modular architecture of BMCs. More broadly, this study has implications for deciphering how bacterial cells modulate the construction and functions of diverse metabolic modules within a single cellular context and could inform the rational design and engineering of synthetic organelles and bio-factories with tailored metabolic functions for biotechnological applications.