MBP Squared

Multiple Sclerosis is a neurological disease affecting around 100,000 people in the UK. The myelin sheath, which insulates nerve fibres, gets broken down – this means that nerve impulses cannot be transmitted.

One of the major proteins in the myelin sheath is Myelin Basic Protein (MBP). It is an unusual protein, it lacks a defined 3-D structure and so it can ‘fall apart’ quite easily. If this happens, then the myelin sheath is attacked by an auto-immune response. One possibility is that phosphorylation of the MBP (adding a phosphate group to specific amino acids) triggers the unfolding of the protein.

We will be looking at how the human MBP protein behaves in the yeast Saccharomyces cerevisiae. This is normal brewer’s and baker’s yeast, but its biochemistry is startlingly similar to humans. We will see how the protein is ‘processed’ by the yeast and try to identify which pathways are involved. This will be possible in yeast since there are entire libraries of strains each of which has one single non-essential gene knocked-out and this represents an enormously powerful tool in dissecting the biochemistry involved.

Details of Research

The students have been divided into six groups, each of which is working on one particular aspect of the project.

  1. E. coli growth and transformation :- this team will grow E. coli and treat it so that it is able to take-up plasmids (small rings of DNA). They will also use the bacteria to bulk-up DNA samples
  2. S. cerevisiae growth and transformation :- this team will do the same as group 1, but in yeast. They will maintain stocks and prepare samples for other groups
  3. Protein purification and analysis :- students will purify proteins using column chromatography and analyse the products by SDS-PAGE (protein gel electrophoresis).
  4. Western blotting :- after transferring proteins form an SDS-PAGE gel onto a nylon membrane, this group will probe the membrane with anti-bodies to identify specific phosphorylation events.
  5. DNA purification and analysis :- students will purify plasmid DNA, cut it with restriction enzymes, analyse the results by agarose gels and create new plasmids ready for transformation.
  6. Bioinformatics :- the “techies” will search data-bases to find similarities between yeast and humans and determine what might be expected from the experiments – they will direct future experiments.