Showing papers in "Applied Biochemistry and Biotechnology in 1990"
TL;DR: Once applied on exudating or bloody tissues, this biofilm displays several advantages as a biological dressing, and hence, it is valuable as a temporary skin substitute in the treatment of skin wounds, such as burns, ulcers, grafts, and as an adjuvant in dermal abrasions.
Abstract: A bacterial strain with morphological and biochemical properties close to Acetobacter xylinum has been cultured in nonagitated, inverted sucrose- and yeast water-based medium for the production of thick, smooth, and floating cellulosic pellicles. The cellulose content (greater than 90%, dry weight, depending on the efficiency of water washing) and the beta-D-homopolyglucan nature of these pellicles were assessed by physical, chemical, and enzymatic methods. The apyrogenic bacterial biomass, a minor component of the dried biofilm (BioFill), is inactivated by ethylene dioxide. Once applied on exudating or bloody tissues, this biofilm displays several advantages as a biological dressing, and hence, it is valuable as a temporary skin substitute in the treatment of skin wounds, such as burns, ulcers, grafts, and as an adjuvant in dermal abrasions.
428 citations
TL;DR: In this paper, the authors present an extension of this concept that produces phenomenological descriptions that permit comparisons of different fractionation proceses and yield some pseudokinetic parameters for aspen acid-catalyzed fractionation.
Abstract: Fractionation of lignocellulosic materials into high-yield cellulosic solid components can be accomplished with acid-catalyzed aqueous and mixed aqueous and nonaqueous media, with the latter leading to high lignin and xyland removal. Concepts combining only time and temperature pulping profile with catalyst concentration are presented to correlate these two types of fractionation methods with pulping variables. Severity concepts combining time and temperature only are often used in the pulp and paper industry. The present extension of this concept producesphenomenological descriptions that permit comparisons of different proceses and yield some pseudokinetic parameters for aspen (Populus tremuloides) acid-catalyzed fractionation.
371 citations
TL;DR: In this article, the microalgae are used to extract storage lipids in large quantitites and thrive in saline waters, and these lipids can then be converted into usable fuel.
Abstract: Microalgae are unique photosynthetic organisms that are known to accumulate storage lipids in large quantitites and thrive in saline waters. Before these storage lipids can be used, they must be extracted from the microalgae and converted into usable fuel. Transesterification of lipids produces fatty acid methyl esters that can be used as a diesel fuel substitute.
231 citations
164 citations
TL;DR: A strategy for immobilization-stabilization of penicillin G acylase from E.coli by multipoint covalent attachment to agarose (aldehyde) gels is developed, able to prepare very active and very stable PGA derivatives containing up to 200 International Units of catalytic activity per mL.
Abstract: We have developed a strategy for immobilization-stabilization of penicillin G acylase from E. coli, PGA, by multipoint covalent attachment to agarose (aldehyde) gels. We hve studied the role of three main variables that control the intensity of these enzyme-support multiinteraction processes: 1. surface density of aldehyde groups in the activated support; 2. temperature; and 3. contact-time between the immobilized enzyme and the activated support prior to borohydride reduction of the derivatives. Different combinations of these three variables have been tested to prepare a number of PGA-agarose derivatives. All these derivatives preserve 100% of catalytic activity corresponding to the soluble enzyme that has been immobilized but they show very different stability. The less stable derivative has exactly the same thermal stability of soluble penicillin G acylase and the most stable one is approximately 1,400 fold more stable. A similar increase in the stability of the enzyme against the deleterious effect of organic solvents was also observed. On the other hand, the agarose aldehyde gels present a very great capacity to immobilize enzymes through multipoint covalent attachment. In this way, we have been able to prepare very active and very stable PGA derivatives containing up to 200 International Units of catalytic activity per mL. of derivative with 100% yields in the overall immobilization procedure.
145 citations
TL;DR: In this paper, the most promising candidates for efficient production of biomass in the US are short rotation hardwoods and herbaceous crops, and two short rotation grasses (weeping lovegrass and switchgrass) and one herbaceous legume (sericea lespedeza) were also obtained from the same source.
Abstract: Large-scale production of ethanol or other transportation fuels by biological conversion of lignocellulosic biomass will eventually require integration with large-scale production of biomass substrates. The most promising candidates for efficient production of biomass in the US are short rotation hardwoods and herbaceous crops. The following samples of short rotation hardwoods were provided by the Biomass Production Program managed for the Department of Energy Biofuels and Municipal Waste Technology Division by the Oak Ridge National Laboratory: Poplar hybrid NE388(Populus, maximowiczii xP. trichocarpa), Poplar hybrid Nil (Populus. trichocarpa xP. deltoides), and Sweetgum(Liquidambar styraciflua). Samples of two short rotation grasses (weeping lovegrass and switchgrass) and one herbaceous legume (sericea lespedeza) were also obtained from the same source. Milled and debarked hardwoods and herbaceous samples were subjected to prehydrolysis with dilute sulfuric acid at 140 and 160 °C for reaction times ranging from 5 to 60 min. The dilute sulfuric acid hydrolyzed all hemicelluloses at longer reaction times (30–60 min at 140 °C and 5–10 min at 160 °C), but solubilized very small amounts (< 15%) of lignin and cellulose. Cellulose in all three pretreated hardwoods became highly digestible by cellulase enzyme fromTrichoderma reesei. The two grasses also responded to dilute acid pretreatment very well.
144 citations
TL;DR: In this paper, the authors compare the model with experimental data and conclude that more detailed studies of the adsorption dynamics, not just the equilibrium, are needed for a more detailed analysis of the model.
Abstract: Comparison of the model with experimental data is currently in progress. It appears that more detailed studies of the adsorption dynamics, not just adsorption equilibrium, are needed.
138 citations
TL;DR: It is reported that yeast xylose and xylulose fermentation can be further improved by cloning and overexpression of thexylulokinase gene, and the resulting genetically-engineered yeasts can ferment xylOSE and xelulose more than twice as fast as the parent yeast.
Abstract: d-Xylose is a major constituent of hemicellulose, which makes up 20–30% of renewable biomass in nature.d-Xylose can be fermented by most yeasts, includingSaccharomyces cerevisiae, by a two-stage process. In this process, xylose is first converted to xylulose in vitro by the enzyme xylose (glucose) isomerase, and the latter sugar is then fermented by yeast to ethanol. With the availability of an inexpensive source of xylose isomerase produced by recombinantE. coli, this process of fermenting xylose to ethanol can become quite effective. In this paper, we report that yeast xylose and xylulose fermentation can be further improved by cloning and overexpression of the xylulokinase gene. For instance, the level of xylulokinase activity in S.cerevisiae can be increased 230fold by cloning its xylulokinase gene on a high copy-number plasmid, coupled with fusion of the gene with an effective promoter. The resulting genetically-engineered yeasts can ferment xylose and xylulose more than twice as fast as the parent yeast.
93 citations
TL;DR: The thermodynamics of 10 industrially-important, enzyme-catalyzed reactions are examined and the available thermodynamic properties, such as equilibrium constants, Gibbs energies, enthalphies, and heat capacity changes are reviewed and summarized.
Abstract: The thermodynamics of 10 industrially-important, enzyme-catalyzed reactions are examined. The reactions discussed are: the conversions of penicillin G to 6-amino-penicillinic acid using the enzyme penicillin acylase; starch to glucose using amylases; glucose to fructose using glucose (xylose) isomerase; cellulose to glucose using cellulase; fumaric acid and ammonia to L-aspartic acid using L-aspartase; transcinnamic acid and ammonia to L-phenylalanine using L-phenylalanine ammonia lyase; L-histidine to urocanic acid and ammonia using L-histidine ammonia lyase; lactose to glucose and galactose using lactase; and the reactions catalyzed by amino acylases and proteases. The selection of these processes was based on the economic value of the products and their intrinsic industrial importance. The available thermodynamic properties, such as equilibrium constants, Gibbs energies (delta G degrees), enthalphies (delta H degrees), and heat capacity changes (delta Cp degrees) of these enzyme-catalyzed reactions, are reviewed and summarized. Recommendations are made for future research in this area.
67 citations
TL;DR: In this article, the authors summarized research to optimize the performance of some of these cultures, and discussed experimental results and models for several reactor designs, including CSTR and packed columns.
Abstract: Liquid and gaseous fuels may be produced biologically from coal by the indirect conversion of coal synthesis gas. Methane has been produced from synthesis gas using acetate and CO2/H2 as intermediates, utilizing a number of CO-utilizing and methanogenic bacteria. Also, a bacterium that is capable of producing ethanol from synthesis gas through indirect liquefaction has been isolated fron natural inocula. This paper summarizes research to optimize the performance of some of these cultures. These conversions, involving H2 and CO, which are only slightly soluble in the liquid media, may be mass transfer limited, and methods to enhance mass transport are examined. Experimental results and models for several reactor designs, including CSTR and packed columns, are presented and discussed.
65 citations
TL;DR: In this paper, the design and application of aqueous two-phase systems for recovery of proteins is reviewed, and the properties of polymers and salt forming the systems and their influence on phase separation and partition are discussed.
Abstract: The design and application of aqueous two-phase systems for recovery of proteins is reviewed. Reference is made to the properties of polymers and salt forming the systems and their influence on phase separation and partition. The properties of systems important for the design of purification strategies are discussed in relation to the surface properties of proteins. Strategies for the modification of systems for bioaffinity partition are considered including the choice of carrier in terms of molecular type and properties, or by particulate addition. Finally, the scaleup of partition is considered, and the reasons for currently limited adoption at larger scales are elucidated.
TL;DR: In this paper, five key issues are identified and considered in relation to the status of current and foreseeable technology, including conversion and production energy balances, suitability of ethanol as a transportation fuel, air quality impacts, raw material supply, and cost.
Abstract: Ethanol produced from lignocellulose is considered as a largescale transportation fuel in the United States. Five key issues are identified and considered in relation to the status of current and foreseeable technology. These are: conversion and production energy balances, suitability of ethanol as a transportation fuel, air quality impacts, raw material supply, and cost. Energy balances and fuel characteristics appear to be consistent with large-scale transportation fuel use of ethanol produced from lignocellulose. Local and global air-quality benefits are expected to accompany use of lignocellulose ethanol. Raw material availability is examined for wastes and for trees and grasses grown as energy crops. Ethanol production levels appear unlikely to be limited by raw material availability as long as economic and other factors are sufficiently favorable to justify allocation of land for this use. Projected ethanol production costs based on current directions of research would allow neat ethanol to become competitive with gasoline by the year 2000 according to current oil price predictions. Biological process steps have the largest contribution to overall costs, are among the least developed aspects of the technology, and appear to have the greatest potential for improvement. Research priorities are discussed.
TL;DR: In this article, the economics of using Na2CO3 rather than CaCO3 as the neutralizing agent in fumaric acid production byRhizopus arrhizus is investigated and compared with the benzene route.
Abstract: The economics of using Na2CO3 rather than CaCO3 as the neutralizing agent in fumaric acid production byRhizopus arrhizus is investigated and compared with the benzene route. Because sodium fumarate, unlike calcium fumarate, is soluble, downstream processing is simplified by requiring less equipment and no heat, and also allows the reuse of cells. In spite of a fumaric acid productivity (g/L/h) 2.4 x lower than the CaCO3 case, the Na2CO3 alternative with cell reuse has a higher rate of return. However, with the current cost of petroleum, the selling price of fumaric acid from the benzene route is half as much as from an idealized Na2CO3 fermentation.
TL;DR: Monoclonal antibodies against carboxypeptidase A and horse radish peroxidase (HRP) were prepared, and those that did not inhibit the respective enzymatic activities were selected.
Abstract: Amino and hydrazyno derivatives of Eupergit C were prepared by reaction of the beads with hexamethylene diamine (HMD) and adipic acid dihydrazide (ADH), respectively. Monoclonal antibodies (mAbs) against carboxypeptidase A (CPA) and horse radish peroxidase (HRP) were prepared, and those that did not inhibit the respective enzymatic activities were selected. The carbohydrate moieties of these antibodies were oxidized by reaction with sodium periodate and then coupled onto the modified beads. The oxidation and coupling reactions were optimized to achieve highly active matrix-conjugated antibodies. Thus, antibody-matrix conjugates that possessed antigen-binding activities close to the theoretical value of 2 mol antigen bound/mol immobilized antibody were obtained.
TL;DR: In semibatch operation with immobilized cells, reproducible results up to 60 cycles were obtained at 50 °C and this operation resulted in no loss of activity of immobilization cells, although the composition of fructo-oligosaccharides was found to be different.
Abstract: Aureobasidium pullulans cells with fructosyltransferase activity were immobilized in 2% calcium alginate beads, and the production of fructo-oligosaccharides from sucrose was studied in a stirred tank bioreactor. It was found that cells ofA. pullulans were entrapped evenly on the alginate matrix of 2.2 mm in diameter, and an effectiveness factor of the beads was determined to be 0.3. By comparison with the system of free cells in batch operation, the total amount of fructo-oligosaccharides produced by immobilized cells was similar although the composition of fructo-oligosaccharides was found to be different. In semibatch operation with immobilized cells, reproducible results up to 60 cycles were obtained at 50 °C and this operation resulted in no loss of activity of immobilized cells.
TL;DR: At low pH values, direct, steady-state fermentation of carbon monoxide to alcohols has been verified and of major significance is the production of butanol from Carbon monoxide in pure culture, which provides a potential mechanism for direct bioconversion of synthesis gas to butanol.
Abstract: Continuous, steady-state fermentations using carbon monoxide gas as the sole carbon and energy source have been achieved with the CO strain ofButyribacterium methylotrophicum. Fermentation pH was found to regulate carbon monoxide metabolism over the pH range of 6.8 to 5.0. Cell growth diminished at low pH, with washout occurring at pH 5.0. As observed previously in batch culture, lower pH values favored production of butyrate over acetate. The mechanism responsible for this trend is currently being investigated by quantification of key intracellular enzyme activities. At low pH values, direct, steady-state fermentation of carbon monoxide to alcohols has been verified. Of major significance is the production of butanol from carbon monoxide in pure culture. This newly identified pathway provides a potential mechanism for direct bioconversion of synthesis gas to butanol.
TL;DR: In this article, mixed-culture studies employing the photosynthetic bacterium R. rubrum for converting CO to CO2 and H2 by the water gas shift reaction and two methanogens, M. formicicum and M. barkeri, for conversion CO 2 and H 2 into CH4 are presented for triculture operation in two types of reactors, the packed bubble column and the trickle-bed reactor.
Abstract: The components of synthesis gas, CO, H2, and CO2, may be converted into CH4 biologically through either acetate or H2/CO2 as intermediates. Of these two routes, conversion through H2/CO2 is preferred. This paper presents results of mixed-culture studies employing the photosynthetic bacteriumR. rubrum for converting CO to CO2 and H2 by the water gas shift reaction and two methanogens,M. formicicum andM. barkeri, for converting CO2 and H2 into CH4. Results are presented for triculture operation in two types of reactors, the packed bubble column and the trickle-bed reactor.
TL;DR: The partition behavior of proteins, nucleic acids, cell membranes, cell organelles and whole cells has been studied in liquid-liquid two-phase systems composed of water, poly-(ethylene glycol), and an hydroxypropyl starch.
Abstract: The partition behavior of proteins, nucleic acids, cell membranes, cell organelles and whole cells has been studied in liquid-liquid two-phase systems composed of water, poly-(ethylene glycol), and an hydroxypropyl starch. The properties of the systems are in many respects comparable with the traditional poly(ethylene glycol)-dextran systems, but the cost is reduced to around one-fifth.
TL;DR: Lipases from different origins have been immobilized in supports chosen by its different aquaphilicity and used as biocatalysts for the hydrolysis of tributyrin and Maximal yields of dibutyrin were found with the solubleCandida lipase, whereas the highest yield was obtained with the least aquaphylic derivative (Candida-Celite).
Abstract: Lipases from different origins have been immobilized in supports chosen by its different aquaphilicity and used as biocatalysts for the hydrolysis of tributyrin. The changes of the concentration of tri-, di-, monobutyrin, glycerol, and butyric acid during the reactions catalyzed by soluble, as well as immobilized, lipases were evaluated by gas chromatography. The experimental data were fitted to a simple kinetic model for the sequential reaction of tributyrin hydrolysis. The calculated apparent rate constants were different for the biocatalysts used and were apparently related to diffusional effects and aquaphilicity of the supports. Maximal yields of dibutyrin were found with the solubleCandida lipase, whereas the highest yield of monobutyrin (90%) was obtained with the least aquaphylic derivative (Candida-Celite).
TL;DR: The ultimate goal of the analytical approaches discussed is to firmly establish structure and, thus, permit the study of structure-function relationships and eventually to allow the intelligent application of carbohydrate remodeling techniques in the preparation of new glycoproteins.
Abstract: Many of the products prepared by biotechnological approaches, including recombinant genetic engineering, cell tissue culture, and monoclonal technologies, are glycoproteins. As little as five years ago, glycosylation was believed to play no significant role in the function of glycoproteins. Recent large scale testing of glycoprotein-based pharmaceuticals has indicated that both the extent and type of glycosylation can play a central role in glycoprotein activity. Although methods for compositional and sequence analysis of proteins and nucleic acids are generally available, similar methods have yet to be developed for carbohydrate oligomers and polymers. This review focuses on new, developing methods for the analysis and sequencing of the carbohydrate portion of glycoproteins. Included are: (1) the release of oligosaccharides and hydrolysis of carbohydrate chains using enzymatic and chemical methods; (2) fractionation by LPLC, electrophoresis, HPLC, and lectin affinity chromatography; (3) detection through the preparation of derivatives or by new electrochemical methods; (4) analysis by spectroscopic methods, including MS and high-field NMR; and (5) their sequencing through the use of multiple, well-integrated techniques. The ultimate goal of the analytical approaches discussed is to firmly establish structure and, thus, permit the study of structure-function relationships and eventually to allow the intelligent application of carbohydrate remodeling techniques in the preparation of new glycoproteins.
Journal Article•
TL;DR: In this paper, the performance and characteristics of a 4.5 m3 nonmixed, vertical flow Experimental anaerobic digestion Test Unit (ETU) treating refuse-derived fuel (RDF) and primary sludge were reported.
Abstract: This paper reports on the performance and characteristics of a 4.5 m3 nonmixed, vertical flow Experimental anaerobic digestion Test Unit (ETU) treating refuse-derived fuel (RDF) and primary sludge. It was operated at 35°C and a loading rate of 3.2 kg VS/m3/d. Three tests involving chemical nutrients addition or effluent supernatant recycle were completed. Hydraulic retention times were 16, 18, and 19 d, respectively. Methane yields ranged from 0.24 to 0.28 m3/kg VS fed, which were 10–20% higher than that from a continuously-stirred tank reactor. Methane compositions were around 58%. Solids distribution, pH and volatile fatty acids (VFA) concentrations profiles, and the ability of microbial communities at various depths to utilize acetate and other VFAs were used to characterize the ETU.
TL;DR: A microbubble dispersion was used to supply oxygen for aerobic fermentations in a standard 2 L stirred tank fermenter and growth rates ofSaccharomyces cerevisiae cultures were found to be equal or greater with MBD sparging than with gas sparged.
Abstract: A microbubble dispersion (MBD) was used to supply oxygen for aerobic fermentations in a standard 2 L stirred tank fermenter. The microbubble dispersion was formed using only surfactants produced naturally. Growth rates ofSaccharomyces cerevisiae cultures were found to be equal or greater with MBD sparging than with gas sparging. The oxygen transfer coefficent with MBD sparging was found to be 190/h and independent of impeller speed from 100–580 rpm. The oxygen transfer coefficient with air sparging rose from 55 to 132/h over the same range of impeller speeds. Power requirements for the fermenter systems were estimated.
TL;DR: In this article, the authors compared the performance of several CO-utilizing and methanogenic bacteria in converting CO, CO2, and H2 to products in the presence of various concentrations of H2S and COS.
Abstract: Anaerobic bacteria have been shown to be capable of converting CO, H2, and CO2 in synthesis gas to valuable products, such as acetate, methane, and ethanol. However, synthesis gas also contains small quantities of sulfur gases such as H2S and COS, that may inhibit the performance of these organisms. This paper compares the performance of several CO-utilizing and methanogenic bacteria in converting CO, CO2, and H2 to products in the presence of various concentrations of H2S and COS. The sulfur gas toxicity levels, growth, substrate uptake, and product formation for each organism are compared.
TL;DR: The highest γ-linolenic acid content was obtained when this alga was cultured under the radiation of around 100 μEinstein/m2/s, which is about one and a half times that of conventional Spirulina platensis.
Abstract: The production of γ-linolenic acid from algae in fresh and marine water was investigated. WhenSpirulina platensis was left in the dark condition, it contained about one and a half times γ-linolenic acid compared with conventionalSpirulina platensis. Marine microalga,Chlorella sp. NKG 042401 contained about 10% of γ-linolenic acid. The highest γ-linolenic acid content was obtained when this alga was cultured under the radiation of around 100 μEinstein/m2/s.
TL;DR: Enzyme immobilization is economically feasible, compared with a system with free enzymes, despite a very high cost for the enzyme attachment, according to a computer program for preliminary cost estimates.
Abstract: A computer program for preliminary cost estimates of free and immobilized enzyme systems has been developed The cost for the hydrolysis of lactose by β-galactosidase fromAspergillus oryzae has been calculated for a batch tank reactor, with free (BTRF) and immobilized (BTRI) enzymes, a continuously stirred tank reactor (CSTR) and a plug-flow tubular reactor (PFTR), considering the mass transfer behavior and deactivation of the enzyme
TL;DR: Among forty strains of marine cyanobacteria isolated in the authors' laboratory, five strains had 1–3 different plasmids, and a hybrid plasmid pUSY02 containing the 1.4kb HindIII fragment of pSY11 andEscherichia coli plasmID pUC18 was constructed, rapidly introduced into marineSynechococcus sp.
Abstract: Among forty strains of marine cyanobacteria isolated in our laboratory, five strains had 1–3 different plasmids. The unicellular marine cyanobacterium,Synechococcus sp. NKBG 042902, contains at least three plasmids (pSY09, pSY10, and pSY11). However, these plasmids are cryptic. Therefore, a hybrid plasmid pUSY02 containing the 1.4kb HindIII fragment of pSY11 andEscherichia coli plasmid pUC18 was constructed. The plasmid pUSY02 transformed both marineSynechococcus sp. NKBG042902-YG1116, which is a cured strain, and fresh waterAnacystis nidulans R2 by dark incubation or Ca2+ treatment. However, the plasmid pSG111 constructed from the plasmid DNA ofA. nidulans R2 failed to transform marineSynechococcus sp. Electroporation was also applicable to transformation of marineSynechococcus sp. and fresh waterA. nidulans R2. The plasmid pUSY02 was rapidly introduced into marineSynechococcus sp.
TL;DR: To test the possible contribution of mycelia-bound enzyme to SSF performance with modern fungal cellulase producers, the performance of SSF using whole culture broth was compared toSSF performance using culture filtrate, and measurements of cellulase activities showed that the broth had more activity than thefiltrate.
Abstract: Cellulase, an enzyme that catalyzes the breakdown of cellulose into glucose, is produced inside fungal cells and secreted into the surrounding media. However, the cells retain significant amounts of β-glucosidase, and this extra enzyme could improve the performance of a simultaneous saccharification and fermentation (SSF) process. To test the possible contribution of mycelia-bound enzyme to SSF performance with modern fungal cellulase producers, the performance of SSF using whole culture broth (with cells) was compared to SSF performance using culture filtrate. Cellulase broth was produced using the advancedTrichoderma reesei mutant L27. Measurements of cellulase activities showed that the broth had more activity than the filtrate. Subsequent SSF experiments were conducted at 37°C in shake flasks withSaccharomyces cerevisiae, and the broth produced 8–25% higher ethanol yields than the cellulase filtrate produced.
TL;DR: In this paper, the performance and characteristics of a 4.5 m3 nonmixed, vertical flow Experimental anaerobic digestion Test Unit (ETU) treating refuse-derived fuel (RDF) and primary sludge were reported.
Abstract: This paper reports on the performance and characteristics of a 4.5 m3 nonmixed, vertical flow Experimental anaerobic digestion Test Unit (ETU) treating refuse-derived fuel (RDF) and primary sludge. It was operated at 35°C and a loading rate of 3.2 kg VS/m3/d. Three tests involving chemical nutrients addition or effluent supernatant recycle were completed. Hydraulic retention times were 16, 18, and 19 d, respectively. Methane yields ranged from 0.24 to 0.28 m3/kg VS fed, which were 10–20% higher than that from a continuously-stirred tank reactor. Methane compositions were around 58%. Solids distribution, pH and volatile fatty acids (VFA) concentrations profiles, and the ability of microbial communities at various depths to utilize acetate and other VFAs were used to characterize the ETU.
TL;DR: Xylitol dehydrogenase from xylose-grown cells of Candida shehatae was purified 215-fold by sequential chromatography on NAD-C8 affinity, Superose-12, and Cibacron blue columns, and a single band was observed by SDS gel electrophoresis, indicating that it was composed of two subunits.
Abstract: Xylitol dehydrogenase (EC1.1.1.9) from xylose-grown cells ofCandida shehatae was purified 215-fold by sequential chromatography on NAD-C8 affinity, Superose-12, and Cibacron blue columns, and a single band was observed by SDS gel electrophoresis. The purified enzyme had a native molecular weight of 82 kDa and a denatured molecular weight of 40 kDa following SDS gel electrophoresis, indicating that it was composed of two subunits. Alcohol dehydrogenase copurified on the NAD-C8 but was substantially removed by Superose-12 and was not detected following Cibacron blue chromatography. The kinetic properties of the C.shehatae xylitol dehydrogenase differed considerably from those described previously for thePachysolen tannophilus enzyme. The Km of the C.shehatae enzyme for xylitol was 3.8 times smaller, whereas the Km for xylulose was 1.7-fold bigger. These factors could account for the lower xylitol production by C.shehatae.
TL;DR: Two biotechnological systems were developed for sucrose conversion into levan and ethanol with Zymomonas mobilis, ensuring a 66.7% transfer of substrate carbon in a batch and 61% carbon transfer in a continuous culture.
Abstract: Two biotechnological systems were developed for sucrose conversion into levan and ethanol withZymomonas mobilis, ensuring a 66.7% transfer of substrate carbon in a batch and 61% carbon transfer in a continuous culture. The effect of glucose, ethanol, and medium pH on sucrose conversion byZ. mobilis was studied. The addition of ethanol to the fermentation medium, in the final conc. of 100 g/L, uncoupled levan synthesis from ethanol fermentation. For a continuous culture, the most efficient conversion of substrate carbon into levan was reached at pH 4.8, giving 64.2 g/L levan, with the levan yield of 0.22 g/g and the productivity of 3.2 g/L/h.