Charlea Clarke

Profile

Project Title: Structural characterization of cobalamin (vitamin B12) biosynthetic enzymes and scavenging proteins

Summary: The focus of this project is to provide structure and mechanistic detail on the components of the B12 uptake and utilization system in commensal bacteria and use this information for biotechnical and medical applications. B12 is an essential cofactor for many enzymes integral for life. However, only one-third of bacterial species can completely synthesize B12 due the enormous metabolic input required for synthesis. Therefore, there is a reliance upon B12 uptake mechanisms in bacteria. These uptake mechanisms utilize specialized outer-membrane lipoproteins that bind B12 with high affinity, this specific binding has useful applications in commercial B12 purification.

Bacteroides thematicron (Bt) is an impressive example of B12  uptake, due to its three conserved genetic loci dedicated to producing B12  binding and transport proteins. Each protein within these loci contribute to the uptake mechanisms that have allowed Bt to colonize the mammalian gut, outcompeting other bacteria as well as the host for this essential vitamin. Therefore, using Bt as a model we will purify these key proteins and structurally characterize their interaction with B12 using various techniques including X-ray crystallography.

Supervisor:

• Prof Richard Pickersgill

Research