Novel tools for bioengineering gut anaerobes

Supervisors: Dr Karen Scott, Dr Petra Louis, Mr Ben Bradley

Project description:

Background

Bioengineering is widely used to incorporate new activities into specific bacteria, for example the industrial production of 1,3-propanediol from glycerol by Clostridium butyricum (1). Obligately anaerobic bacteria resident in the human gut produce a myriad of metabolites of value to the biotechnology industry including organic fatty acids, alcohol and antimicrobials. Modified gut bacteria could overproduce these metabolites for commercial use, or could be used as biotherapeutics to provide prolonged and controlled in situ delivery of drugs/metabolites into the human intestine.

In this project, we address the urgent need to find new host microbes to produce industrial biochemicals by mining the human gut microbiota and screening for unique genes and robust strains. Important chemical targets are the chiral chemicals 3-hydroxybutyrate (3-HB) and 1,3-butanediol (1,3-BDO), as well as the biofuel butanol. Redesigning the metabolism of butyrate-producing Clostridium species, to produce 3-HB and 1,3-BDO or butanol, creates a microbial producer of high value, speciality chemicals.

Research strategy

Objective 1 Creation of a new series of modular plasmids. The extensive, unique collection of human gut bacteria held at the Rowett will be mined for plasmid replication origins and antibiotic resistance genes. These genes will be fully characterised and tested for efficacy following introduction into the existing pMTL80000 series of modular plasmid vectors provided by the partner company CHAIN (2).

Objective 2 The host range and stability of the new series of modular plasmids will be determined by mating with a broad phylogenetic range of gut anaerobic bacteria from the RINH bacterial collection, and additional commercial bacteria known to grow effectively in large scale production vessels.

Objective 3 Plasmids developed in Objectives 1 and 2 will be engineered to contain new genes targeting the butyrate synthesis pathway, to produce the end-products 3-hydroxybutyrate and 1,3-butanediol. These plasmids will be transformed into robust, fast-growing butyrate producing strains, and the altered metabolite synthesis confirmed. Genes for the secondary conversion of butyrate to butanol will also be cloned into plasmids which will be used to engineer butyrate producing gut bacteria to overproduce the biofuel butanol.

Objective 4 Fermentation conditions will be adapted to optimise production of the novel chemicals from the engineered strains. During this objective the student will gain experience in quality control required for industrial manufacturing and production upscaling.

Research Training:

The student would receive training in anaerobic culturing, metabolic engineering, advanced DNA transfer techniques, bioinformatics, and controlled continuous fermentation techniques. The lab at the Rowett has extensive experience of working with anaerobic intestinal bacteria, and maintains a diverse collection of key human gut bacterial isolates.

The associated company, CHAIN Biotechnology Ltd. is focussed on creating new biotechnologies capable of unlocking the potential of a large range of Gram-positive organisms. The company will provide business training for the student aligned with the project theme. Working within the company provides a valuable insight into commercially focused operations and a better understanding on translating research outputs to customer need. This placement will fulfil the PIPS training of the student.

References

  1. Gonzalez-Pajuelo, M. et al (2005) Metabolic engineering of Clostridium acetobutylicum for the industrial production of 1,3-propanediol from glycerol. Metabolic Engineering 7: 329-336.

Heap, J.T.  et al (2009) A modular system for Clostridium shuttle plasmids J. Microbiol. Methods. 78: 79-85.

Other: