Extended operation of a pressurized 75-L bioreactor for shLkn-1 production by Pichia pastoris using dissolved oxygen profile control.
In this study, a dissolved oxygen (DO)-stat fed-batch process was conducted in a pressurized 75-L bioreactor, resulting in the production of the short version of human leukotactin-1 (shLkn-1) using Pichia pastoris as the host, with control of the DO-stat profile and an extension of the recombinant shLkn-1 production phase. By regulation of the exhaust-gas valve, we were able to maintain the vessel pressure at up to 120 kPa, in order to overcome DO limitations associated with the use of the DO-stat. The lowest DO value was adjusted by varying the feed pump speed, allowing us to control the DO-stat profile. This principle was successfully applied to both glycerol feeding during the growth phase and methanol feeding for the induction of shLkn-1. The extension of the methanol induction phase to a total of 192 h of culture time resulted in a shLkn-1 concentration of 2.5 g/L, and a total of 102 g of cumulative production. During this extended induction period, the C-terminal residue of shLkn-1 was truncated and this was confirmed by both reverse-phase HPLC and mass spectrometry.
Woo
Sung-Hwan
SH
Mogam Biotechnology Institute, Yougin, Kyonggi-Do, Korea.
Park
See-Hyoung
SH
Lim
Hyung-Kwon
HK
Jung
Kyung-Hwan
KH
D019149:Bioreactors; D019742:Chemokines, CC / Q000096:biosynthesis*; D007218:Industrial Microbiology / Q000379:methods*; D019402:Macrophage Inflammatory Proteins; D000432:Methanol / Q000378:metabolism; D015846:Monokines / Q000096:biosynthesis*; D010100:Oxygen; D010843:Pichia / Q000254:growth & development / Q000378:metabolism*; D011312:Pressure; D011994:Recombinant Proteins / Q000096:biosynthesis; D012995:Solubility
C108814:CCL15 protein, human; D019742:Chemokines, CC; D019402:Macrophage Inflammatory Proteins; D015846:Monokines; D011994:Recombinant Proteins; D010100:Oxygen; D000432:Methanol