Maximization of Candida utilis cytochromes by pulse additions of ethanol to continuous cultures

Candida utilis was grown with glucose as growth-limiting carbon source in batch culture, steady-state continuous culture, and non-steady-state continuous culture. High cytochrome concentrations were routinely recorded in cells harvested in the latter stages of batch culture. They were occasionally recorded in cells from imprecisely controlled steady-state cultures, but precise control of the steady-state dissolved oxygen tension stabilized the cytochromes at relatively low levels. Controlled non-steady-state continuous cultures, imposed by pulse additions of ethanol, routinely produced cells with high cytochrome concentrations. A mechanism is proposed whereby maintenance of cytochrome derepression in continuous culture is dependent upon indefinitely prolonging an “overshoot” response in gene expression.     

The development of granulation in an upflow floc digester and an upflow anaerobic sludge blanket digester treating cane juice stillage

Summary The use of synthetic polyelectrolytes in the Upflow Floc digester during the treatment of high strength cane juice stillage, resulted in a more rapid accumulation of biomass and promoted granule formation at an earlier stage compared to a control upflow anaerobic sludge blanket digester (UASB). In the Upflow Floc reactor the granules were composed of rod shaped organisms, whereas in the UASB the granules were primarily filamentous. Both types of granules had good settling properties and high activities.     

Anaerobic digestion of high-strength cheese whey utilizing semicontinuous digesters and chemical flocculant addition

Semicontinuous digesters were used to anaerobically treat high-strength whey (70 kg/m(3).COD). A maximum loading of 16.1 kg COD/m(3).day was obtained with soluble COD removal efficiencies greater than 99%. The use of a chemical flocculant resulted in an increased biomass concentration in the digester compared to a control, thus enabling correspondingly higher space loadings to be applied. With the onset of substantial levels of granulation of the biomass, flocculant dosage was able to be discontinued. This article discusses the performance of the digesters in detail and, briefly, the long-term operational difficulties experienced and the control strategies employed on such systems.     

Increased a-lytic protease production from recombinant Escherichia coli grown with oxygen limitation

Production of a mutant of the enzyme a-lytic protease from recombinant E. coli JM109 increased from 50 units/l to 750 units/l as the dissolved oxygen was decreased from above 30% saturation to near zero during batch growth on a complex medium. This represents a 60-fold increase in specific protease production (units/g cell dry wt.). Limiting the oxygen supply decreased the maximum growth rate from 0.55 h -1 to 0.066 h -1 which may in turn regulate the synthesis of recombinant protease in this system so that increased levels of product are obtained.     

Kinetic analysis of L-lysine production by a fluoropyruvate sensitive mutant of B.lactofermentum

The kinetics of L-lysine production by a fluoropyruvate sensitive (FP^s) mutant of B.lactofermentum have been analysed in batch and continuous culture. Under optimal conditions (viz. T = 30° C, pH = 7.0, DO = 10 % air saturation) it was established that the initial phosphate level could be used to extend the batch culture to give an L-lysine.HCl concentration of 68.0 g/l and a yield of 0.43 g/g. In continuous culture the yield increased at low dilution rates under conditions similar to those for extended batch culture.     

Cytoskeletal modulation of lipid interactions regulates Lck kinase activity

The actin cytoskeleton promotes clustering of proteins associated with cholesterol-dependent rafts, but its effect on lipid interactions that form and maintain rafts is not understood. We addressed this question by determining the effect of disrupting the cytoskeleton on co-clustering of dihexadecyl-(C(16))-anchored DiO and DiI, which co-enrich in ordered lipid environments such as rafts. Co-clustering was assayed by fluorescence resonance energy transfer (FRET) in labeled T cells, where rafts function in the phosphoregulation of the Src family kinase Lck. Our results show that probe co-clustering was sensitive to depolymerization of actin filaments with latrunculin B (Lat B), inhibition of myosin II with blebbistatin, and treatment with neomycin to sequester phosphatidylinositol 4,5-bisphosphate. Cytoskeletal effects on lipid interactions were not restricted to order-preferring label because co-clustering of C(16)-anchored DiO with didodecyl (C(12))-anchored DiI, which favors disordered lipids, was also reduced by Lat B and blebbistatin. Furthermore, conditions that disrupted probe co-clustering resulted in activation of Lck. These data show that the cytoskeleton globally modulates lipid interactions in the plasma membrane, and this property maintains rafts that function in Lck regulation.     

Behavior of the hybrid plasmid pNSW301 in Zymomonas mobilis grown in continuous culture

The stability of the plasmid pNSW301 which was formed by cointegration of the Inc W R plasmid Sa and the 14.5-kb pNSW1 plasmid of Zymomonas mobilis ZM6100 was investigated in ZM6100(pNSW301) grown in continuous culture without antibiotic selection. The cointegrate plasmid, pNSW301, was found to be structurally unstable and a total reduction in the size of pNSW301 of approximately 21 kb occurred during growth in continuous culture. Following a systematic study, a number of deletion derivatives of pNSW301 were isolated and used to transform Escherichia coli HB101, with the exception of pNSW312. The plasmid pNSW312 was 100% stable in ZM6100(pNSW312) in continuous culture but was unable to replicate in E. coli.     

Cold shock response inLactococcus lactis ssp.diacetylactis

The acquired freeze-thaw tolerance was investigated forLactococcus lactis ssp.diacetylactis. Pretreatment of microorganisms at less severe temperatures to initiate cold tolerance gaveL. lactis ssp.diacetylactis improved cell viability after successive freezings and thawings. The ability of cells to survive freeze-thaw was dependent on factors experienced prior to freezing. Factors affecting lactic acid bacteria survival during freeze-thaw cycles were found to be different diluents, growth phase, and different cold temperatures. Viability experiments showed that this strain displaying cold shock cryotolerance had an improved survival capacity in stationary phase. The plasmid contents of lactic acid bacteria isolated from different types, DRC-2 and DRC-2C, were examined and compared with the plasmid contents of culture collection strains both before and after cold shock treatment. Using agarose gel electrophoresis, no obvious correlation between the cold shock response and the number of plasmids in the cell could be observed.