Team:Tec Guadalajara/Molecular Biology
The pathway from glucose to IA is already known in Penicillium. It consists of two steps catalyzed by two enzymes.
- Oxidation of D-glucose to D-glucono-1,5-lactone by glucose oxidase (GOX)
- Oxidation of D-gluconolactone to D-isoascorbic acid by D-gluconolactone oxidase (GLO)
Both genes were optimized for Pichia pastoris and synthesized. Our chassis was methylotrophic yeast Pichia pastoris GS115.
- No P. pastoris strains to overcome codon bias
- Codon optimization strategies are usually based on Kazusa's codon usage database
- We used the COOL web tool using Bai et al's (2011) codon table based on highly expressed genes
Advantages of biological reduction of GO
The production of isoascorbic acid by using P. pastoris allows us to:
- Reduce GO without compromising operator's health and environment
- Scale-up the process. However, it must be improved to be economically feasible
- Show proof of concept to move from a chemical to a biological process