The Biotransformation of Propylene to Propylene Oxide by Methylococcus capsulatus (Bath): 3. Reactivation of Inactivated Whole Cells to Give a High Productivity System

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Abstract
Methylococcus capsulatus (Bath) possesses methane monooxygenases (soluble - (sMMO) and particulate - (pMMO)) which are able to catalyse the epoxidation of propylene to propylene oxide. In a previous paper we have shown that the production of the epoxide caused a rapid inactivation of the bioconversion process (Stanley et al, 1992). This paper shows that cultures containing pMMO, inactivated by propylene oxide production, could be completely reactivated in the presence of growth substrates within 5 h after the removal of propylene oxide so long as the propylene oxide production rate was below 150 nmol min−1 [mg dry weight cells]−1. Reactivation under these conditions was detectable within 30 min of propylene oxide removal. On the other hand, cells inactivated by propylene oxide production rates in excess of 150 nmol min−1 [mg dry weight]−1 did not begin to recover activity within the 30 min period. Furthermore a lag period was observed before reactivation began which was dependent upon the initial production rate. Cultures possessing sMMO took twice as long to recover their activity compared with cells containing pMMO. Reactivation of propylene oxide production could occur without growth, but the process required the presence of a carbon and energy source (methane or methanol), sulphur, nitrogen and oxygen, although copper (which is normally involved in pMMO activity) was not required. It was shown that de novo protein synthesis was required for reactivation of activity. Production rates of 12 g 1−1 d−1 could be maintained for longer than three weeks in a single phase production process and rates up to 30 g 1−1 d−1 were achieved in a two stage process. Using Methylocystis parvus (OBBP) rates of up to 90 g 1−1 d−1 were attained over a one week period.