Enzymes in a box: Engineering bacterial microcompartments for biotechnological applications

BMCs are proteinaceous partitions that sequester metabolic pathways from the rest of the cytosolic pool. Several BMC types have been characterized and shown to prevent cell toxicity from toxic intermediates by quickly metabolizing them to non-toxic products (termed as metabolosomes) or to enhance CO2 fixation by preventing its diffusion outside of the BMC (termed as carboxysomes). To achieve the aforementioned activities, BMCs enclose several enzymes (termed as core enzymes) in a large quasi-icosahedral proteinaceous shell of about 100-400 nm in size. The shell of the BMCs is formed by hexameric, pentameric and trimeric proteins, termed as shell proteins, which have been purified and crystallized successfully, revealing pores in the centre of their polymeric structures. These protein pores (4-14 Å) facilitate (passive) metabolite transport likely in a charge-dependent way. Thus, metabolites enter the BMC through the pores of the shell proteins and are efficiently converted in the lumen of BMCs by encapsulated core enzymes.

Background

BMCs have triggered the interest of many international research groups for fundamental but also applied research. A central application of BMCs is for metabolic engineering where BMCs can serve as modular “enzymes in a box”. By keeping the reaction-relevant enzymes in close proximity, BMCs can (presumably) facilitate enhanced metabolisms by overcoming slow reaction rates, resource competition, co-factor imbalance, by-product formation or metabolite toxicity.

Aim of the project

We want to re-purpose BMCs and turn them into nanofactories. What we can do with this technology is virtually limitless. As such, we want to discover how far we can push this technology. The initial aim of this project is to be able to encapsulate target proteins in the lumen of BMCs and produce target metabolites. We also want to discover the physical properties of BMCs at the single molecule level. To do this we collaborate with Dr. Sonja Schmid  of the Biophysics department and with specialists on mass spectrometry and structural biochemistry at Utrecht University.

Techniques

Multiple techniques are used to achieve our targets including CRISPR-Cas, Advanced microscopy, Anaerobic and aerobic fermentation, HPLC, GC, FPLC and regular plasmid design and cloning. Handling of multiple microorganisms is also possible.

• See all microbiology thesis projects