Sunday, April 29, 2007
2-13

Factors influencing expression of the ligninolytic enzyme manganese peroxidase

Christine J. Kelly1, Curtis A. Lajoie1, and Fei Jiang2. (1) Chemical Engineering, Oregon State University, 103 Gleeson Hall, Corvallis, OR 97333, (2) Biomedical and Chemical Engineering, Syracuse University, Link Hall, Syracuse, NY 13244

The effects of pH and temperature on recombinant manganese peroxidase (rMnP) production by the yeast Pichia pastoris αMnP1-1 were investigated in shake-flask and fed-batch fermentations. P. pastoris αMnP1-1 constitutively expresses the manganese peroxidase (mnp1) cDNA from Phanerochaete chrysosporium, and the rMnP has similar kinetic characteristics and pH activity as the wild-type MnP (wtMnP). Cultivation of P. chrysosporium mycelia in stationary flasks for production of wt-heme peroxidases is commonly conducted at low pH (pH 4.2) (Tien and Kirk, ). Experiments with P. pastoris αMnP1-1 demonstrated that rMnP production is highest at pH 6, with rMnP concentrations in the medium declining rapidly at pH less than 5.5, even though cell growth rates were similar from pH 4-7. Reducing the pH of the fermentation from 6 to 4.5 resulted in a rapid decline in rMnP activity, and spiking of pH 4.5 fermentations with concentrated rMnP also yielded rapid declines in enzyme activity. rMnP stability tests with bioreactor cultures and culture supernatants indicated that rMnP is less stable in pH 4.5 culture supernatant than in supernatant samples from pH 6 fermentations, the presence of yeast cells in the stability test increases the rate of loss of rMnP activity, samples taken later in the fermentation are more against rMnP than samples taken earlier, and lowering the temperature from 30oC to 20oC decreases the rate of loss of activity. These results are consistent with the hypothesis that intracellular proteases are released from dead and lysed cells that are active against rMnP at pH less than 5.5.

 


See more of Poster session 1
See more of General Submissions

See more of The 29th Symposium on Biotechnology for Fuels and Chemicals (April 29 - May 2, 2007)