Bacterial microcompartments are protein-based organelles that can enclose enzymes
and other proteins, however without successful isolation of the compartments they do
not have much use. We used methods previously described [1] to isolate the
microcompartments through lysis and ultracentrifugation then confirmed isolation of the
micro compartments in two ways:
1. Fluorescent foci (Fusion Red-LVA and Fusion Red Luciferase LVA) to identify the
compartments in fluorescent imaging. With Eut-C tags, the fluorescent markers
accumulate inside the compartment, allowing them to be clearly visible inside the
compartment. The LVA protein degradation tag targets any excess fluorescent proteins
for degradation to allow a sufficient background for viewing the compartments in vivo.
2. Electron Microscopy with various radiological stains on a copper grid. We found that flash freezing the compartments in liquid ethane before viewing with electron microscopy preserved the integrity of the compartment.
Figure 1: EutS compartment containing fusion red
tagged with Eut C
2. Electron Microscopy with various
radiological stains on a copper grid.
Figure 2: EutS compartment with negative stain