Vitamin B12 into Yeast
Professor Martin Warren, University of Kent
“This project gave me valuable lab experiences…. I explored techniques such as cloning, protein expression, purification, microbiology and imaging. Carrying out most of the work individually boosted my confidence level going into my final year project and also for future research.”
Alice Le, Undergraduate Student, University of Kent
The project was initiated to test whether yeasts and fungi have the ability to take up cobalamin (also known as vitamin B12), even though they do not use this molecule for any known biochemical process. If yeasts are able to absorb this nutrient, then it might be possible to extend the metabolic repertoire of reactions that can be performed in this host.
The CBMNet Vacation Scholarship was used to investigate three aspects of vitamin B12 in yeast. Firstly, we investigated if yeasts could absorb vitamin B12. We undertook bioassays to determine how much B12 was absorbed by cultures of S. cerevisiae, C. albicans and S. pombe that had been grown in increasing concentrations of exogenous cobalamin.
Secondly, we wanted to visualize B12 uptake through the use of live cell imaging of yeasts that had been grown in the presence of fluorescent derivatives of B12. Finally, we wanted to see if B12 levels in S. cerevisiae could be increased through the expression of a recombinant B12 binding protein (BtuF).
We were successful in demonstrating that several different yeasts have the ability to absorb B12 from the growth media. This uptake become saturated, suggesting that the uptake process involves a specific transporter.
The B12 uptake could be followed using fluorescence microscopy, but detailed sub-cellular analysis of the distribution of the nutrient was not undertaken. Although the plasmid containing BtuF was transformed into S. cerevisiae, the yeast did produce any recombinant protein.
Our results suggest that yeasts could be engineered to harbour B12-dependent processes in the future.
First we will repeat some experiments, especially with a view to producing a B12-binding protein within the yeast cell. This will complete the information we need to demonstrate unequivocally that yeasts can absorb B12 and that proteins within the cytoplasm can bind the nutrient. Longer term, we are in touch with several companies who are interested in engineering yeasts so that they expand the metabolic range of the organism.
“The project yielded some exciting results. This research holds significant potential to expand the metabolic capabilities of yeast – and thereby allow the organism to make altogether new compounds.”
Prof Martin Warren, University of Kent