A Major Shell Protein of 1,2‐Propanediol Utilization Microcompartment Conserves the Activity of Its Signature Enzyme at Higher Temperatures

Abstract

A classic example of an all‐protein natural nano‐bioreactor, the bacterial microcompartment is a prokaryotic organelle that confines enzymes in a small volume enveloped by an outer protein shell. These protein compartments metabolize specific organic molecules, allowing bacteria to survive in restricted nutrient environments. In this work, 1,2‐propanediol utilization microcompartment (PduMCP) was used as a model to study the effect of molecular confinement on the stability and catalytic activity of native enzymes in the microcompartment. A combination of enzyme assays, spectroscopic techniques, binding assays, and computational analysis were used to evaluate the impact of the major shell protein PduBB′ on the stability and activity of PduMCP′s signature enzyme, dioldehydratase PduCDE. While free PduCDE shows ∼45 % reduction in its optimum activity (activity at 37 °C) when exposed to a temperature of 45 °C, it retains similar activity up to 50 °C when encapsulated within PduMCP. PduBB′, a major component of the outer shell of PduMCP, preserves the catalytic efficiency of PduCDE under thermal stress and prevents temperature‐induced unfolding and aggregation of PduCDE in vitro. We observed that while both PduB and PduB′ interact with the enzyme with micromolar affinity, only the PduBB′ combination influences its activity and stability, highlighting the importance of the unique PduBB′ combination in the functioning of PduMCP.

Innate and engineered attributes of bacterial microcompartments for applications in bio-materials science

“…3) showed that the major shell protein of Pdu BMC shows This journal is © The Royal Society of Chemistry 2023 chaperone-like activity by conserving the structure and activity of its signature enzyme under thermal stress. 51 2.2.2. Catalytic machineries inside the protein envelopeenzymatic core.…”

“…Further, the protein shell also protects the encapsulated enzymatic cargo with high thermal and pH stability. 51 Another important property of bacterial microcompartments is that they can self-assemble heterogeneously, and each shell protein can self-assemble, homogenously forming different nanostructures based on the concentration in both in vivo and in vitro environments. All these features of BMCs, the high catalytic efficiency, multifunctionality, semi-permeability and self-assembling nature of the protein shell make BMCs superior to other protein compartments for exploitation as biosynthetic nano-reactors.…”

“…106 An advantage of BMCs as drug delivery vehicles is that they can shelter the cargo from thermal and pH stress. 51,107 However, the biocompatibility of BMC needs to be explored.…”

“…3) showed that the major shell protein of Pdu BMC shows chaperone-like activity by conserving the structure and activity of its signature enzyme under thermal stress. 51…”

“…Fig.3The chaperone-like activity of the PduBB 0 shell protein of Salmonella enterica on the enzyme, PduCDE, providing thermal stability to the enzyme. (Adapted from Kumar et al51 ).…”