摘要:

AbstractOn‐line determination of expression rates from cloned genes inEscherichia coliand of plasmid copy number would be useful for monitoring accumulation of non‐secreted proteins. As an initial model for monitoring gene expression in intact cells, a non‐gene‐fusion enzyme‐based indicator plasmid has been constructed containing thephoAgene coding for alkaline phosphatase (AP) in pUCIS and pACYC184. The activity of AP can be rapidly determined in permeabilized cells. A flow injection analysis (FIA) assay has been developed which allows the direct real‐time measurement of the AP activity during cell growth. A model target gene coding forE. colicyanase (cynS) has been inserted in order to determine the ratio between the expression of the target and indicator, AP. A linear relationship has been found between plasmid copy number and AP activity for the high‐copy pUC vector. To minimize indicator expression, transcription terminators have been inserted between thecynSandphoAgenes, altering the target‐to‐indicator ratio by 10‐ to 40‐fold. These vectors may be useful for the rapid continuous determination of plasmid copy number and target gene expression for nonsecreted proteins and would overcome the limitations ofin situprobe biosensors for real‐time determination of the accumulation of proteins from

ISSN:0006-3592    

DOI:10.1002/bit.260340802

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractGramicidin S synthetase, the enzyme complex catalyzing the biosynthesis of the antibiotic gramicidin S inBacillus brevis, is subject to O2‐dependentin vivoinactivation during exponential aerobic growth after reaching a peak in specific activity. The five amino acid substrates of the synthetase are capable of stabilizing its activity to varying degrees in whole cells shaken aerobically. Depending on the time of cell harvesting before, during, or after the peak in intracellular gramicidin S synthetase specific activity, the enzyme has a long, medium, or short half‐life, respectively. The kinetic profiles of gramicidin S synthetase inB. breviscells indicate that both the kinetics of synthetase loss and the degree of its amino‐acid‐mediated stabilization are a strong function of the cells' physiological deve

ISSN:0006-3592    

DOI:10.1002/bit.260340803

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractIn continuous aerobic treatment of artificial sewage by activated sludge, the rate of removal of chemical oxygen demand (COD) was markedly enhanced by the presence of cross‐linked poly‐4‐vinylpyridine (PVP). The concentration of dissolved oxygen (DO) was also low in the presence of PVP. The extent of improvement in COD removal increased with increases in substrate load and the surface area of the PVP in the working space of the test apparatus. These results suggest an increase of the bacterial population resulting from the presence of the PVP. However, formation of measurable biofilm was not detected on the surface of the PVP during continuous aerobic trea

ISSN:0006-3592    

DOI:10.1002/bit.260340804

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractPyrococcus furiosusrepresents one of the most important hyperthermophilic bacteria isolated thus far because of its relatively high cell yields and rapid growth rates.Pyrococcus furiosusexhibits several interesting growth characteristics, especially in terms of biotic gas production, which were examined in this study. In the presence of elemental sulfur, both carbon dioxide and hydrogen sulfide production appeared to be strongly growth associated, while no significant hydrogen production was observed. In the absence of sulfur, hydrogen and carbon dioxide were produced by the organism and hydrogen inhibition was observed. The addition of elemental sulfur to the medium apparently eliminated, hydrogen inhibition as growth proceeded normally even when hydrogen was added to the gas phase. Also, no apparent substrate limitation or toxic product could be attributed to the cessation of growth as cell growth in spent media was at least as good as in fresh media. An unstructured growth model was used to correlate growth and gas production forP. furiosusin complex seawater‐based media at 98° C both in the absence and presence of elemental sulfur. The model was shown to be useful for examining some of the observations made in this stu

ISSN:0006-3592    

DOI:10.1002/bit.260340805

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractThe effect of oxygen supply on the growth of suspension cultures ofCatharanthus roseusin Erlenmeyer flasks was investigated. Below a critical oxygen supply rate the culture could not survive. By increasing the oxygen supply, a point is reached where the culture survives but no growth is possible. At higher oxygen supply rates there is a regime where both growth rate and the maximum biomass concentration increase with oxygen supply. Eventually there comes a point where no further increase in biomass is achieved, probably due to the depletion of the sugars; however, the growth rate continues to increase with oxygen supply until a maximum growth rate is obtained. The ratio of fresh to dry weight at maximum fresh weight increased with shaker table speed of rotation accompanied by a greater rate of sugar depletion.

ISSN:0006-3592    

DOI:10.1002/bit.260340806

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractClostridium formicoaceticumhomofermentatively converts lactate to acetate at 37°C and pH 6.6–9.6. However, this fermentation is strongly inhibited by acetic acid at acidic pH. The specific growth rate of this organism decreased from a maximum at pH 7.6 to zero at pH 6.6. This inhibition effect was found to be attributed to both H+and undissociated acetic acid. At pH values below 7.6, the H+inhibited the fermentation following non‐competitive inhibition kinetics. The acetic acid inhibition was found to be stronger at a lower medium pH. At pH 6.45–6.8, cell growth was found to be primarily limited by a maximum undissociated acetic acid concentration of 0.358 g/L (6mM). This indicates that the undissociated acid, not the dissociated acid, is the major acid inhibitor. At pH 7.6 or higher, this organism could tolerate acetate concentrations of higher than 0.8M, but salt (Na+) became a strong inhibitor at concentrations of higher than 0.4M. Acetic acid inhibition also can be represented by noncompetitive inhibition kinetics. A mathematical model for this homoacetic fermentation was also developed. This model can be used to simulate batch fermentation at any pH between 6.9 a

ISSN:0006-3592    

DOI:10.1002/bit.260340807

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractA microfiltration cell‐recycle pilot‐scale system was developed comprised of a conventional continuous‐flow fermentor connected to anin situsteam‐sterilizable cross‐flow ceramic filter with a backflushing device. A microcomputer was used to control filtration pressure, tangential flow velocity, and backflushing. Performance of the system was tested with the anaerobic production of thermostable extracellular β‐amylase at 60°C byClostridium thermosulfurogeneson maltose or malto‐dextrin media. Filtration rates during continuous cultivation were between 20 and 60 L/m2/h. The maltodextrin and cell debris occurring at high retentate flow rates or filtration pressures impaired the performance of the filter. Backflushing initially improved the permeate flux to 42% in a maltose medium and to 10% in a maltodextrin medium, but the effect diminished with time. The productivity of β‐amylase (as much as 48 U/mL/h) and concentration of biomass (as much as 14 g/L) were increased 11‐ and 12‐fold, respectively, if compared to values obtained in a chemostat. The concentration of β‐amylase rose to 220 U/mL in the reactor, which was 5.5‐fold more than under comparabl

ISSN:0006-3592    

DOI:10.1002/bit.260340808

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractThe desire to increase cell density and product concentration has been the primary driving force for the development of better animal cell culture processes. In the technique used in our laboratory—microencapsulation—insect cells (Spodoptera frugiperda), infected with a temperature‐sensitive mutant of theAutographa californicanuclear polyhedrosis virus (AcNPV), were cultured in multiple membrane alginate–polylysine (PLL) microcapsules which had a controlled membrane molecular‐weight cutoff and an intracapsular alginate concentration which was ca. 16% lower than that obtained in the commercially available single‐membrane system. Cell culture experiments indicated that the intracapsular alginate concentration appears to be a key factor in achieving good cell growth. It was possible to obtain intracapsular cell densities of 8 × 107cells/mL capsules and virus concentrations to 109IFU/mL capsules. The virus litre in the supernatant was ca. 300 times lower, indicating that virtually all of the virus was retained within

ISSN:0006-3592    

DOI:10.1002/bit.260340809

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractCellulase was covalently immobilized on an enteric coating polymer,Eudragit L, that is reversibly soluble and insoluble depending on the pH of the medium. The hydrolysis of solid cellulose with the immobilized enzyme can take advantage of the soluble property of the immobilized enzyme itself at the most reactive pH value; on the other hand, recovery of the enzyme can take advantage of the insoluble property of the enzyme at other pH values. It was experimentally confirmed that 100% of immobilized enzyme activity in solution can be recovered by precipitation and by dissolving it again by alternative change of pH. After a period of hydrolysis, immobilized enzyme and unreacted cellulose were precipitated together to remove the product—the soluble sugar solution—by changing pH. Following this, a new buffer solution was added to the precipitate to dissolve it and resume the reaction. This was repeated several times. The hydrolysis rate of this process increased significantly compared with that of a batch process. Utilization of the reversible soluble–insoluble carrier for immobilizing enzyme is promising, not only for cellulose–cellulase systems, but also for other heterogeneous reaction

ISSN:0006-3592    

DOI:10.1002/bit.260340810

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractA new isolation and purification method for bioproducts using membrane‐encapsulated affinity adsorbents was investigated. The new method involves encapsulation of affinity adsorbents, batch adsorption of the bioproduct from whole fermentation broth and rapid batch desorption after dissolution of the capsule membranes. Recovery of protein A fromStaphylococcus aureuswas used as the model experimental system. Affinity adsorbents such as rabbit IgG‐agarose were successfully encapsulated within calcium alginate membranes and used directly to recover protein A from whole cell homogenate containing a number of macromolecular contaminants as well as suspended solids. Both high yield and high purity of protein A were recovered by this method in comparison with various previously reported meth

ISSN:0006-3592    

DOI:10.1002/bit.260340811

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY