Alginate formation in Azotobacter vinelandii UWD during stationary phase and the turnover of poly-beta-hydroxybutyrate

Azotobacter vinelandii UWD is a mutant of strain UW that is defective in th e respiratory oxidation of NADH. This mutation causes an overproduction of polyhydroxyalkanoates (PHAs), as polyester synthesis is used as an alternat ive electron sink. Since PHAs have potential for use as natural, biodegrada ble plastics, studies of physiology related to their production are of inte rest. Alginate production by this strain is limited to <11 <mu>g (mg cell p rotein)(-1) which permits high efficiency conversion of carbon source into PHA. However, greater than or equal to 400 mug (mg cell protein)(-1) was fo rmed when UWD cells were oxygen-limited and in the stationary phase of grow th. Alginate formation was fuelled by PHA turnover, which was coincident wi th the synthesis of alkyl resorcinols, under conditions of exogenous glucos e limitation. However, alginate production was a phenotypic and reversible change. Alginate production was stopped by interruption of algD with Tn5lac Z. LacZ activity in UWD was shown to increase in stationary phase, while La cZ activity in a similarly constructed mutant of strain UW did not. Transcr iption of algD in strain UWD started from a previously identified RpoD prom oter and not from the AlgU (RpoE) promoter. This is because strain UWD has a natural insertion element in algU. Differences between strain UW and UWD may reside in the defective respiratory oxidation of NADH, where the NADH s urplus in strain UWD may act as a signal of stationary phase. Indeed, a bac kcross of UW DNA into UWD generated NADH-oxidase-proficient cells that fail ed to form alginate in stationary phase. Evidence is also presented to show that the RpoD promoter may be recognized by the stationary phase sigma fac tor (RpoS), which may mediate alginate production in strain UWD.