Effects of agitation intensity on mycelial morphology and protein production in chemostat cultures of recombinant Aspergillus oryzae

Abstract

Abstract:The effects of agitation on fragmentation of a recombinant strain of Aspergillus oryzae and its consequential effects on protein production have been investigated. Constant mass, 5.3-L chemostat cultures at a dilution rate of 0.05 h −1 and a dissolved oxygen level of 75% air saturation, have been conducted at 550, 700, and 1000 rpm. These agitation speeds were chosen to cover a range of specific power inputs (2.2 to 12 kW m −3 ) from realistic industrial levels to much higher values. The use of a constant mass chemostat linked to a gas blender allowed variation of agitation speed and hence gas holdup without affecting the dilution rate or the concentration of dissolved oxygen. The morphology of both the freely dispersed mycelia and clumps was characterized using image analysis. Statistical analysis showed that it was possible to obtain steady states with respect to morphology. The mean projected area at each steady state under growing conditions correlated well with the ''energy dissipation/circulation'' function, [P/(kD 3 t c )], where P is the power input, D the impeller diameter, t c the mean circulation time, and k is a geometric constant for a given impeller. Rapid transients of morphological parameters in response to a speed change from 1000 to 550 rpm probably resulted from aggregation. Protein production (␣-amylase and amyloglucosidase) was found to be independent of agitation speed in the range 550 to 1000 rpm (P/V = 2.2 and 12.6 kW m −3 , respectively), although significant changes in mycelial morphology could be measured for similar changes in agitation conditions. This suggests that mycelial morphology does not directly affect protein production (at a constant dilution rate and, therefore, specific growth rate). An understanding of how agitation affects mycelial morphology and productivity would be valuable in optimizing the design and operation of large-scale fungal fermentations for the production of recombinant proteins.

Suitable extracellular oxidoreduction potential inhibit

“…As a result, part of NADH was oxidized by H 2 O 2 that resulted in the lower NADH/NAD + ratios under oxidative condition. During the fermentation of Actinomycetes, high stirring speed damages the mycelium [18]. And the mycelium morphology of Actinomycetes plays an important role in polyketides production [19].…”

Agitation effects on morphology and protein productive fractions of filamentous and pelleted growth forms of recombinant Aspergillus niger

“…On the other hand, production of an extracellular cutinase by recombinant Aspergillus awamori was found to be independent on the agitation intensity in fed-batch cultures [25]. Also, production of extracellular amyloglucosidase by recombinant Aspergillus oryzae was reported to be independent on power input variations in chemostat cultures at constant dilution rate [38]. During batch cultivation, however, production of extracellular amyloglucosidase increased with increasing agitation speed while agitation speed variations had no significant effect on protein production in fed-batch cultures [39].…”

“…In particular, power input or agitation intensity has been identified as an important variable influencing the fungal morphology and productivity in recombinant Aspergillus protein production processes [25,28,[37][38][39]. In general, increasing power input changes the growth morphology from pelleted to filamentous growth forms.…”

Optimization of Medium Composition and Cultivation Parameters for Fructosyltransferase Production by Penicillium aurantiogriseum AUMC 5605

“…On the other hand, oxygen dependence at the lower agitation rate varied with the substrate type and its concentration (Lejeune and Baron, 1995). In much fungal fermentation, the high apparent viscosities and the nonNewtonian behavior of the broths necessitate the use of high agitation speeds to provide adequate mixing and oxygen transfer (Amanullah et al, 1999). However, mycelia damage at higher stirring speeds can limit the acceptable range of speeds.…”