Biosynthesis And Structural Characterization of Levan By A Recombinant Levansucrase From Bacillus Subtilis ZW019

Abstract Purpose: The yield of levan extracted from microbial fermentation broth is low, so in vitro catalytic synthesis of levan by levansucrase is expected to be one of the industrial production approaches of levan. Methods: A recombinant plasmid pET-28a-AcmA-Z constructed in the previous study was used to produce levansucrase. The effects of temperature, pH, and metal ions on the levan formation activity of the levansucrase were investigated. The polymer was analyzed by means of HPIC, FTIR, NMR techniques.Results: The recombinant levansucrase could be easily purified in one step and the purified enzyme had a single band clearly visible in SDS-PAGE. The conditions for enzymatic reactions was optimal at pH 5.2 and 40 ℃, and the activity of enzymes was stimulated by K+ and Ca2+. The yield of levan biosynthesis from 10% (w/v) sucrose with 6.45 U/g sucrose of levansucrase was 30.6 g/L. The molecular weight of the levan was about 1.56×106 Da, as measured by GPC. HPIC analysis showed that the monosaccharide composition of the levan was fructose and glucose. The results of FTIR and NMR analysis indicated that the polymer produced by the recombinant levansucrase was β-(2, 6) levan.Conclusions: The results of this study provide a basis for large-scale production of levan by enzymatic method.

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Biosynthesis and Structural Characterization of Levan by a Recombinant Levansucrase From Bacillus Subtilis ZW019

10.21203/rs.3.rs-511002/v1 ◽

2021 ◽

Author(s):

Jingyue Wang ◽

Xinan Xu ◽

Fangkun Zhao ◽

Nan Yin ◽

Zhijiang Zhou ◽

...

Keyword(s):

Large Scale ◽

Fermentation Broth ◽

Monosaccharide Composition ◽

Enzymatic Reactions ◽

Scale Production ◽

Large Scale Production ◽

Sds Page ◽

One Step ◽

Activity Of Enzymes

Abstract ObjectivesThe yield of levan extracted from microbial fermentation broth is low, so in vitro catalytic synthesis of levan by levansucrase is expected to be one of the industrial production approaches of levan. ResultsA recombinant plasmid Pet-28A-AcmA-Z constructed in the previous study was used to produce levansucrase. The recombinant levansucrase could be easily purified in one step and the purified enzyme had a single band clearly visible in SDS-PAGE. The conditions for enzymatic reactions was optimal at pH 5.2 and 40 ℃, and the activity of enzymes was stimulated by K+ and Ca2+. The yield of levan biosynthesis from 10% (w/v) sucrose with 6.45 U/g sucrose of levansucrase was 30.6 g/L. The molecular weight of the levan was about 1.56×106 Da, as measured by GPC. HPIC analysis showed that the monosaccharide composition of the levan was fructose and glucose. The results of FTIR and NMR analysis indicated that the polymer produced by the recombinant levansucrase was β-(2, 6) levan.ConclusionsThe results of this study provide a basis for large-scale production of levan by enzymatic method.

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A Review on Additive Manufacturing Possibilities for Electrical Machines

Energies ◽

10.3390/en14071940 ◽

2021 ◽

Vol 14 (7) ◽

pp. 1940

Author(s):

Muhammad Usman Naseer ◽

Ants Kallaste ◽

Bilal Asad ◽

Toomas Vaimann ◽

Anton Rassõlkin

Keyword(s):

Additive Manufacturing ◽

Large Scale ◽

Three Dimensional ◽

Electrical Machines ◽

Electromagnetic Properties ◽

Scale Production ◽

Performance Parameters ◽

Large Scale Production ◽

One Step ◽

Manufacturing Techniques

This paper presents current research trends and prospects of utilizing additive manufacturing (AM) techniques to manufacture electrical machines. Modern-day machine applications require extraordinary performance parameters such as high power-density, integrated functionalities, improved thermal, mechanical & electromagnetic properties. AM offers a higher degree of design flexibility to achieve these performance parameters, which is impossible to realize through conventional manufacturing techniques. AM has a lot to offer in every aspect of machine fabrication, such that from size/weight reduction to the realization of complex geometric designs. However, some practical limitations of existing AM techniques restrict their utilization in large scale production industry. The introduction of three-dimensional asymmetry in machine design is an aspect that can be exploited most with the prevalent level of research in AM. In order to take one step further towards the enablement of large-scale production of AM-built electrical machines, this paper also discusses some machine types which can best utilize existing developments in the field of AM.

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Optimizing in vitro large scale production of giant reed (Arundo donax L.) by liquid medium culture

Biomass and Bioenergy ◽

10.1016/j.biombioe.2014.07.004 ◽

2014 ◽

Vol 69 ◽

pp. 21-27 ◽

Cited By ~ 13

Author(s):

Valeria Cavallaro ◽

Cristina Patanè ◽

Salvatore L. Cosentino ◽

Isabella Di Silvestro ◽

Venera Copani

Keyword(s):

Liquid Medium ◽

Large Scale ◽

Arundo Donax ◽

Giant Reed ◽

Scale Production ◽

Large Scale Production ◽

Medium Culture ◽

Arundo Donax L

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Large-scale production of polyoma middle T antigen by using genetically engineered tumors

Molecular and Cellular Biology ◽

10.1128/mcb.5.7.1795-1799.1985 ◽

1985 ◽

Vol 5 (7) ◽

pp. 1795-1799

Author(s):

D R Kaplan ◽

B Bockus ◽

T M Roberts ◽

J Bolen ◽

M Israel ◽

...

Keyword(s):

Nude Mice ◽

Large Scale ◽

T Antigen ◽

Genetically Engineered ◽

Scale Production ◽

Large Scale Production ◽

Metallothionein Promoter ◽

Polyoma Middle T Antigen ◽

Nih 3T3

A recombinant plasmid containing a metallothionein promoter-polyoma middle T cDNA fusion was constructed and used to transfect NIH 3T3 cells. Transformed cells expressing middle T were injected into nude mice. Within 3 weeks, each mouse produced tumors containing middle T equivalent to that in 250 to 1,000 100-mm dishes of polyomavirus-infected cells. This middle T, partially purified by immunoaffinity chromatography, retained activity as measured by its ability to be phosphorylated in vitro. The combined approach of fusing strong promoters to genes of interest and utilizing nude mice to grow large quantities of cells expressing the gene provides a quick, inexpensive alternative to other expression systems.

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Production of Phytotoxic Metabolite Using Biphasic Fermentation System from Strain C1136 of Lasiodiplodia pseudotheobromae, a Potential Bioherbicidal Agent

Notulae Scientia Biologicae ◽

10.15835/nsb9310082 ◽

2017 ◽

Vol 9 (3) ◽

pp. 371-377

Author(s):

Charles Oluwaseun ADETUNJI ◽

Julius Kola OLOKE ◽

Gandham PRASAD ◽

Moses ABALAKA ◽

Emenike Onyebum IROKANULO

Keyword(s):

Large Scale ◽

Wild Strain ◽

Fermentation Medium ◽

Scale Production ◽

Conventional Farming ◽

Large Scale Production ◽

Phytotoxic Metabolite ◽

Biphasic Fermentation

Formulation of effective and environmental friendly bioherbicides depends on the type of fermentation medium used for the production of phytotoxic metabolites. The effect of biomass, colony forming unit and the phytotoxic metabolite produced from the biphasic fermentation was carried out, while the phytotoxic metabolite was tested in vivo and in-vitro on Echinochola crus-galli and dicotyledonous Chromolaena odorata. The mutant strain of Lasiodiplodia pseudotheobromae C1136 (Lp90) produced the highest amount of conidia and the largest necrotic area on the two tested weeds when compared to its wild strain in the different biphasic media combinations. The study revealed that the biphasic system containing PDB + rice produced the highest bioherbicidal activities. Therefore, the phytotoxic metabolites from strain C1136 are suggested for large scale production of bioherbicides for the management of weeds in conventional farming to improve yield and enhance food security.

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The large-scale production of monoclonal antibodies in vitro

Monoclonal antibodies in biology and biotechnology:theoretical and practical aspects ◽

10.1017/cbo9780511565137.005 ◽

2009 ◽

pp. 265-315

Author(s):

Kenneth C. McCullough ◽

Raymond E. Spier

Keyword(s):

Monoclonal Antibodies ◽

Large Scale ◽

Scale Production ◽

Large Scale Production

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Efficient Production of l-Ribose with a Recombinant Escherichia coli Biocatalyst

Applied and Environmental Microbiology ◽

10.1128/aem.02768-07 ◽

2008 ◽

Vol 74 (10) ◽

pp. 2967-2975 ◽

Cited By ~ 30

Author(s):

Ryan D. Woodyer ◽

Nathan J. Wymer ◽

F. Michael Racine ◽

Shama N. Khan ◽

Badal C. Saha

Keyword(s):

Escherichia Coli ◽

Large Scale ◽

Active Form ◽

Apium Graveolens ◽

Efficient Production ◽

Scale Production ◽

Gene Encoding ◽

Large Scale Production ◽

One Step ◽

Rare Sugars

ABSTRACT A new synthetic platform with potential for the production of several rare sugars, with l-ribose as the model target, is described. The gene encoding the unique NAD-dependent mannitol-1-dehydrogenase (MDH) from Apium graveolens (garden celery) was synthetically constructed for optimal expression in Escherichia coli. This MDH enzyme catalyzes the interconversion of several polyols and their l-sugar counterparts, including the conversion of ribitol to l-ribose. Expression of recombinant MDH in the active form was successfully achieved, and one-step purification was demonstrated. Using the created recombinant E. coli strain as a whole-cell catalyst, the synthetic utility was demonstrated for production of l-ribose, and the system was improved using shaken flask experiments. It was determined that addition of 50 to 500 μM ZnCl2 and addition of 5 g/liter glycerol both improved production. The final levels of conversion achieved were >70% at a concentration of 40 g/liter and >50% at a concentration of 100 g/liter. The best conditions determined were then scaled up to a 1-liter fermentation that resulted in 55% conversion of 100 g/liter ribitol in 72 h, for a volumetric productivity of 17.4 g liter−1 day−1. This system represents a significantly improved method for the large-scale production of l-ribose.

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