Dissolved-oxygen-stat controlling two variables for methanol induction of rGuamerin in Pichia pastoris and its application to repeated fed-batch

Abstract

A DO-stat control strategy for two variables was introduced to the rGuamerin production process in Pichia pastoris and applied to repeated fed-batch culture. Two interrelated variables, namely the ratio of partial pressure of pure O2 in the inlet air-stream and the methanol feed rate, were controlled simultaneously. By using this control strategy, methanol feeding for induction could be controlled automatically while efficiently controlling the dissolved oxygen level. As a result, the cell concentration reached more than 140 g l(-1) and rGuamerin expression level 450 iu l(-1). rGuamerin was secreted into the culture medium and reached a level that was 40% higher than achieved in a fed-batch process using manual control of the methanol feeding rate. Repeated rGuamerin induction was achieved by repeating the methanol feeding and withdrawing the culture broth during extended production. During more than 250 h of culture, expression of rGuamerin was maintained at an average of about 430 iu l(-1 )(473 mg l(-1)), without causing the cell density to decrease. In addition to the rGuamerin production process, the proposed control system might be applied to cultivation of other methylotrophic yeasts in the production of therapeutic proteins.

Bioprocess strategies for mass multiplication of and metabolite synthesis by plant growth promoting pseudomonads for agronomical applications

“…For 23-32 h of cultivation, the culture was subjected to a transition period where the feed was added in pulses for culture adaptation. Here, oscillatory behavior of DO was observed whenever a feed of 5 mL was added manually with a gap period of 2 h. Similar oscillatory behavior in DO profile has been observed by others also [24,25]. During high magnitude oscillations in DO, the feed addition usually increases the chances of substrate accumulation in the bioreactor.…”

Production of monoclonal antibodies by glycoengineered Pichia pastoris

“…Metabolism of methanol in P. pastoris involves the reaction of methanol with oxygen catalyzed by alcohol oxidase to form formaldehyde (Cereghino and Cregg, 2000;Lueers et al, 1998;Ren et al, 2003). The low affinity of alcohol oxidase for oxygen implies that the methanol uptake rate depends on the availability of oxygen (Couderc and Baratti, 1980;Jungo et al, 2007a;Lim et al, 2003;Veenhuis et al, 1983). It is therefore reasonable to postulate that a reduced methanol uptake rate under oxygen limited conditions, resulted in a reduced cellular growth rate that in turn led to a reduction in volumetric productivity.…”

A simple model‐based control for Pichia pastoris allows a more efficient heterologous protein production bioprocess

“…In the first method, the methanol-feeding rate is modified in order to avoid methanol exhaustion as indicated by a sharp increase in dissolved oxygen (Lim et al, 2003;Rodríguez Jiménez et al, 1997).…”