Team:Paris Bettencourt/Project/Phytase

Ferment It Yourself

iGEM Paris-Bettencourt 2O15

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Phytase

Background	Aims	Results
The yeast Saccharomyces cerevisiae naturally produces phytase (like some microoganisms).	Make Idli batter Saccharomyces cerevisiae produce high amount of phytase during fermentation to increase mineral aivability of the batter.	Successfully avoided the 2 repressors genes of the phytase biosynthesis pathway of and in Saccharomyces cerevisiae.

Introduction

Anemia affects one third of the world's population, mostly in relation to iron deficiencies. Anemia and similar mineral deficiency diseases are primarily widespread in developing countries like India, partly resulting from the local diet that is mainly made up of cereal grains and seeds such as rice . In these types of food, iron bioavailability is substantially reduced by the presence of phytic acid (C₆H₁₈O₂₄P₆) that chelates minerals and forms insoluble salts which precludes their absorption in the gastrointestinal tract.
Current research on increasing the bioavailability of iron or zinc involves the bioengineering of crop plants which not only poses challenges in terms of the production of efficient genetically modified crops but also requires extensive research for drawing any conclusions on strain sustainability.
We propose an alternative strategy that focuses on the bioengineering of microorganisms involved in the fermentation of idli, a dish widely used as primary food source throughout much of India. Indeed, the lab model organism Saccharomyces cerevisiae is a strain present in the idli microbiome that naturally produces phytases. Phytases are enzymes that are able to hydrolyze phytic acid even when complexed with minerals, resulting in a greater mineral bioavailability.
However the production of phytases in Saccharomyces cerevisiae is down-regulated by two genes : PHO80, present on chromosome 15 and PHO85, found on chromosome 16. The knockout of these genes would likely increase the yield of phytase production and therefore increase the general bioavailability of minerals in fermatation-based dished such as idli.

Figure 1: Phytic acid in complex with calcium, magnesium, zinc and iron

Figure 2:Phytase hydrolyzes phytic acid.

Experimental design

We used the yeast Saccharomyces cerevisiae, more precisly the strain SK1 coming from the INSERM unit U1001. We create two plasmids to replace the two repressors genes called here PHO80 and PHO85. The plasmid used here is pSB1C3, we design two pairs of primers to take the gene of geneticin resistance and the gene of RFP

Results

Bibliography

Veide, J. & Andlid, T. Improved extracellular phytase activity in Saccharomyces cerevisiae by modifications in the PHO system. International Journal of Food Microbiology 108, 60-67 (2006).