http://compbio.korea.ac.kr/wiki/images/8/83/Mmbr.1996.expression.pdf

Strategies for achieving high-level expression of genes in Escherichia coli.

Bacillus species continue to be dominant bacterial workhorses in microbial fermentations. Bacillus subtilis (natto) is the key microbial participant in the ongoing production of the soya-based trad...

Developments in the use of Bacillus species for industrial production

The term “alkaliphile” is used for microorganisms that grow optimally or very well at pH values above 9 but cannot grow or grow only slowly at the near-neutral pH value of 6.5. Alkaliphiles include prokaryotes, eukaryotes, and archaea. Many different taxa are represented among the alkaliphiles, and some of these have been proposed as new taxa. Alkaliphiles can be isolated from normal environments such as garden soil, although viable counts of alkaliphiles are higher in samples from alkaline environments. The cell surface may play a key role in keeping the intracellular pH value in the range between 7 and 8.5, allowing alkaliphiles to thrive in alkaline environments, although adaptation mechanisms have not yet been clarified. Alkaliphiles have made a great impact in industrial applications. Biological detergents contain alkaline enzymes, such as alkaline cellulases and/or alkaline proteases, that have been produced from alkaliphiles. The current proportion of total world enzyme production destined for the laundry detergent market exceeds 60%. Another important application is the industrial production of cyclodextrin by alkaline cyclomaltodextrin glucanotransferase. This enzyme has reduced the production cost and paved the way for cyclodextrin use in large quantities in foodstuffs, chemicals, and pharmaceuticals. It has also been reported that alkali-treated wood pulp could be biologically bleached by xylanases produced by alkaliphiles. Other applications of various aspects of alkaliphiles are also discussed.

Alkaliphiles: Some Applications of Their Products for Biotechnology

Reliable cloning of functional antibody variable domains from hybridomas and spleen cell repertoires employing a reengineered phage display system

The methylotrophic industrial yeast, Pi–chia pastoris, has been developed as a host system for the large–scale production of heterologous proteins. As an example of the systems, the synthesis of hepatitis B surface antigen (HBsAg) is described. Expression of the HBsAg gene is controlled by a methanol–regulated promoter derived from the primary alcohol oxidase gene, AOX1, of P. pastoris. The highest level of HBsAg is observed in a novel mutant strain of P. pastoris in which the AOX1 structural gene is deleted and replaced by a construction composed of the AOX1 promoter–HBsAg gene expression cassette. When grown on methanol, 2–3% of the soluble protein in this strain is HBsAg. Characterization of the recombinant HBsAg indicates that virtually all of the P. pastoris–synthesized HBsAg exists in a form which is similar to human serum–derived HBsAg 22 nm particles. Scale–up of the HBsAg production process from shake–flask to high cell–density fermentor cultures is described as well. A single culture of the P. pastoris HBsAg expression strain in a volume of 240 liters and at a density of 60 grams/liter dry weight of cells produced 0.4 grams/liter, or a total of 90 grams of HBsAg 22 nm particles, an amount sufficient for approximately 9 million doses of vaccine.

High–Level Expression and Efficient Assembly of Hepatitis B Surface Antigen in the Methylotrophic Yeast, Pichia Pastoris

In order to minimize the adverse effect of CO2 gas in a packed bed immobilized yeast reactor, a fluidized bed reactor was used for the continuous production of ethanol from glucose. Immobilized yeast was prepared by entrapping whole cells of Saccharomyces cerevisiae within a Caalginate matrix. It was found that the efficiency of the ethanol production in a fluidized bed reactor was 100% better than that for a packed bed reactor system. The alcohol productivity obtained was 21 g/l/hr in a fluidized bed reactor at 94% of conversion level.

Continuous ethanol production by immobilized yeast in a fluidized reactor

Immobilised yeast trapped in an alginate matrix demonstrated maximum activity at 30 degrees C and showed no pH effect between 3 and 7. Substrate inhibition was observed at glucose concentrations above 8% but the immobilised cells retained 70% of their maximum activity at 20% glucose concentration. The operation stability of immobilised cells was lower in simple glucose solution than in the activation medium in which only 20% of the activity was lost after 10 days operation. Inactivated immobilised yeast beads were reactivated by incubation in activation medium without a significant increase in cell numbers in a bead. During the operation of the immobilised yeast in a packed bed reactor, CO/sub 2/ gas accumulation adversely affected the reactor performance. An ideal plus flow reactor, not taking into account the formation of CO/sub 2/ gas bubbles and the presence of mass trasnfer resistance, was simulated using a kinetic model for the production of ethanol and the simulation results were compared with the actual reactor performance to determine the CO/sub 2/ gas effect, quantitatively. Up to 45% of the substrate conversion was lost due to the accumulation of CO/sub 2/ gas bubbles in all cases. (Refs. 21).

Ethanol production by immobilised yeast and its CO2 gas effects in a packed bed reactor

A theoretical model equation was derived to find the correlation between the conversion and the amount of immobilized penicillin amidase in column. The theoretical values of the conversion were predicted form this correlation and compared with experimental results. It was observed in a column reactor that the pH drop along the column path was linear versus the enzyme loading and that the enzyme activity was also linearly dependent on pH up to 8.0. In order to diminish the effect of pH drop, a continuous two-stage plug-flow reactor (PFR) with pH adjustment between the two columns was used was used in the experiments, and two- and three-stage PFRs were simulated by computer. In the case of the two-stage PFR, the maximum productivity was demonstrated experimentally and theoretically as well. when an equal amount of the immobilized enzyme was packed in both columns. It was also predicted in the tree-stage PFR system that the optimal distributions of enzyme loading in three columns were found to be 1:1:1. It was demonstrated that the increased number of reactors in series could enhance the level of the maximum productivity with a given amount of enzyme loading.

The production of 6‐aminopenicillanic acid by a multistage tubular reactor packed with immobilized penicillin amidase

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1016/0014-5793%2883%2980337-8

Rifamycin B. oxidase from Monocillium spp., a new type of diphenol oxidase

Cassava and sago starch were evaluated for their feasibilities as substrates for ethanol production using Zymomonas mobilis ZM4 strain. Before fermentation, the starch materials were pretreated employing two commercial enzymes, Termamyl (thermostable α-amylase) and AMG (amyloglucosidase). Using 2 μl/g of Termamyl and 4 μl/g of AMG, effective conversion of both cassava and sago starch into glucose was found with substrate concentration up to 30%(w/v) dry substances. Fermentation study performed using these starch hydrolysates as substrates resulted in ethanol yield at an average of 0.48g/g by Z. Mobilis ZM4.

Moon Hi Han

Papers

Continuous ethanol production by immobilized yeast in a fluidized reactor

Ethanol production by immobilised yeast and its CO2 gas effects in a packed bed reactor

The production of 6‐aminopenicillanic acid by a multistage tubular reactor packed with immobilized penicillin amidase

Rifamycin B. oxidase from Monocillium spp., a new type of diphenol oxidase

Ethanol production from cassava and sago starch using Zymomonas mobilis