Secretion of Kluyveromyces lactis Cu/Zn SOD: strategies for enhanced production

The Kluyveromyces lactis Cu/Zn SOD gene (SOD1) was fused with the toxin K1 signal sequence to obtain extracellular production of superoxide dismutase. Kluyveromyces marxianus L3 and K. lactis MW98-8C strains were transformed and compared as hosts for the secretion. The effects of the media composition were evaluated: In K. lactis, the highest volumetric activity was obtained in YKK synthetic medium in the presence of Cu²⁺/Zn²⁺ cofactors (9.6 kU l⁻¹). In K. marxianus, active SOD was produced only in YPD medium supplemented with Cu²⁺ and Zn²⁺ (8.8 kU l⁻¹). In order to improve the production of secreted active SOD in K. lactis, the SOD1 copper carrier (CCS1) was overexpressed and targeted to the secretory apparatus. A positive effect was observed only when K. lactis was grown in a medium without Cu²⁺/Zn²⁺ supplement. The best performing culture conditions for K. lactis and K. marxianus recombinant strains were successfully applied to two laboratory-scale fed-batch processes, and volumetric SOD activities increased up to 19.4 and 24.1 kU l⁻¹, respectively.