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Industrial Enzymes (industrial + enzyme)
Selected AbstractsPurification and molecular characterization of subtilisin-like alkaline protease BPP-A from Bacillus pumilus strain MS-1LETTERS IN APPLIED MICROBIOLOGY, Issue 3 2006T. Miyaji Abstract Aims:, The present study was conducted by screening zein-degrading bacteria in an attempt to obtain zein-degrading protease. Methods and Results:, Soil bacteria were screened by formation of a clear zone on zein plates. Characterization of a zein-degrading bacterium indicated a taxonomic affiliation to Bacillus pumilus, and was named MS-1 strain. The strain produced two different types of extracellular proteases, BPP-A and BPP-B. In this study, we purified and characterized BPP-A because it exhibited a higher ability to hydrolyze zein than BPP-B. When casein was used as the substrate, the optimal pH for BPP-A was 11·0. In BPP-A, zein was better substrate than casein at pH 13·0, whereas casein was better one than zein at pH 11·0. The bppA gene encoded a 383-amino acid pre-pro form of BPP-A, and mature BPP-A contained 275 amino acid residues. It was concluded that BPP-A belonged to the subtilisin family. Conclusion:, A zein-degrading bacterium assigned to B. pumilus produced two different types of extracellular proteases, BPP-A and BPP-B. BPP-A exhibited an ability to hydrolyze zein in an extreme alkaline condition. Significance and Impact of the Study:, This is a first report on screening for zein-degrading micro-organisms. The subtilisin-like protease BPP-A is possible to utilize as an industrial enzyme for the production of zein hydrolysates. [source] Indian Biotech Bazaar: A swot analysisBIOTECHNOLOGY JOURNAL, Issue 5 2007Abhishek Kumar Biotechnology is a life science-based technique especially used in agriculture, medicine and food sciences. It is generally defined as the manipulation in organisms to generate products for the welfare of the world. Biotechnology combines disciplines such as genetics, biochemistry, microbiology, and cell biology along with information technology, chemical engineering, robotics etc. It includes basic industries such as food processing, tissue culture, plant development and other sophisticated ones such as recombinant therapeutics and diagnostics. Biotechnology, globally recognized as a rapidly emerging and far-reaching technology, is aptly described as the "technology of hope" for its promise of food, health and environmental sustainability. In India, biotechnology employs more than 10 000 people and generates roughly US$ 500 million in revenue annually. The biotechnology market has increased its sales from Rs. 50 billion in 1997 to Rs.70 billion in 2000, and is expected to cross Rs. 240 billion by the year 2010. In India, the human health biotech products account for 60% of the total market; agribiotech and veterinary 25%, medical devices, contract research and development (R&D), reagents and supplies constitute the remaining 15% Moreover, to facilitate foreign investment, capital and government policies are being revised. Other important industries include industrial enzyme manufacture, bioinformatics, and medical devices. Biotechnology has had limited appeal so far on our capital markets, and we have less then a dozen biotech companies listed on the public markets. [source] An in silico method using an epitope motif database for predicting the location of antigenic determinants on proteins in a structural contextJOURNAL OF MOLECULAR RECOGNITION, Issue 1 2006Vincent Batori Abstract Presently X-ray crystallography of protein,antibody complexes is still the most direct way of identifying B-cell epitopes. The objective of this study was to assess the potential of a computer-based epitope mapping tool (EMT) using antigenic amino acid motifs as a fast alternative in a number of applications not requiring detailed information, e.g. development of pharmaceutical proteins, vaccines and industrial enzymes. Using Gal d 4 as a model protein, the EMT was capable of identifying, in the context of the folded protein, amino acid positions known to be involved in antibody binding. The high sensitivity and positive predictive value of the EMT as well as the relevance of the structural associations suggested by the EMT indicated the existence of amino acid motifs that are likely to be involved in antigenic determinants. In addition, differential mapping revealed that sensitivity and positive predictive value were dependent on the minimum relative surface accessibility (RSA) of the amino acids included in the mapping, demonstrating that the EMTs accommodated for the fact that epitopes are three-dimensional entities with various degrees of accessibility. The comparison with existing prediction scales demonstrated the superiority of the EMT with respect to physico-chemical scales. The mapping tool also performed better than the available structural scales, but the significance of the differences remains to be established. Thus, the EMT has the potential of becoming a fast and simple alternative to X-ray crystallography for predicting structural antigenic determinants, if detailed epitope information is not required. Copyright © 2005 John Wiley & Sons, Ltd. [source] Stabilization of proteins by low molecular weight multi-ionsJOURNAL OF PHARMACEUTICAL SCIENCES, Issue 10 2002Donald S. Maclean Abstract A method is described to identify small molecule ligands that stabilize proteins. The procedure is based on the hypothesis that molecules of various sizes containing two to four charges should occasionally bind to unpaired charged sites on the surface of proteins and by crosslinking such residues stabilize the native state of the liganded protein. A simple turbidity assay is employed that detects inhibition of protein aggregation under selected sets of conditions. Eight test proteins were screened and in all cases specific ligands were identified that inhibited protein aggregation at millimolar to micromolar concentrations. Only small effects of these stabilizers on protein biological activities were found. In some, but not all cases, circular dichroism and fluorescence studies provided direct evidence of the binding of stabilizing ligands to the proteins suggesting multiple mechanisms of stabilization. This approach should be applicable to the development of excipients for the stabilization of pharmaceutical proteins and industrial enzymes as well as serve as starting points for second-generation inhibitors of increased affinity and specificity. © 2002 Wiley-Liss Inc. and the American Pharmaceutical Association J Pharm Sci 91:2220,2229, 2002 [source] Approaches to achieve high-level heterologous protein production in plantsPLANT BIOTECHNOLOGY JOURNAL, Issue 1 2007Stephen J. Streatfield Summary Plants offer an alternative to microbial fermentation and animal cell cultures for the production of recombinant proteins. For protein pharmaceuticals, plant systems are inherently safer than native and even recombinant animal sources. In addition, post-translational modifications, such as glycosylation, which cannot be achieved with bacterial fermentation, can be accomplished using plants. The main advantage foreseen for plant systems is reduced production costs. Plants should have a particular advantage for proteins produced in bulk, such as industrial enzymes, for which product pricing is low. In addition, edible plant tissues are well suited to the expression of vaccine antigens and pharmaceuticals for oral delivery. Three approaches have been followed to express recombinant proteins in plants: expression from the plant nuclear genome; expression from the plastid genome; and expression from plant tissues carrying recombinant plant viral sequences. The most important factor in moving plant-produced heterologous proteins from developmental research to commercial products is to ensure competitive production costs, and the best way to achieve this is to boost expression. Thus, considerable research effort has been made to increase the amount of recombinant protein produced in plants. This research includes molecular technologies to increase replication, to boost transcription, to direct transcription in tissues suited for protein accumulation, to stabilize transcripts, to optimize translation, to target proteins to subcellular locations optimal for their accumulation, and to engineer proteins to stabilize them. Other methods include plant breeding to increase transgene copy number and to utilize germplasm suited to protein accumulation. Large-scale commercialization of plant-produced recombinant proteins will require a combination of these technologies. [source] Wet-milling transgenic maize seed for fraction enrichment of recombinant subunit vaccineBIOTECHNOLOGY PROGRESS, Issue 2 2010Lorena Moeller Abstract The production of recombinant proteins in plants continues to be of great interest for prospective large-scale manufacturing of industrial enzymes, nutrition products, and vaccines. This work describes fractionation by wet-milling of transgenic maize expressing the B subunit of the heat-labile enterotoxin of Escherichia coli (LT-B), a potent immunogen and candidate for oral vaccine and vaccine components. The LT-B gene was directed to express in seed by an endosperm specific promoter. Two steeping treatments, traditional steeping (TS, 0.2% SO2 + 0.5% lactic acid) and water steeping (WS, water only), were evaluated to determine effects on recovery of functional LT-B in wet-milled fractions. The overall recovery of the LT-B protein from WS treatment was 1.5-fold greater than that from TS treatment. In both steeping types, LT-B was distributed similarly among the fractions, resulting in enrichment of functional LT-B in fine fiber, coarse fiber and pericarp fractions by concentration factors of 1.5 to 8 relative to the whole kernels on a per-mass basis. Combined with endosperm-specific expression and secretory pathway targeting, wet-milling enables enrichment of high-value recombinant proteins in low-value fractions, such as the fine fiber, and co-utilization of remaining fractions in alternative industrial applications. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2010 [source] Use of Glycol Ethers for Selective Release of Periplasmic Proteins from Gram-Negative BacteriaBIOTECHNOLOGY PROGRESS, Issue 5 2007Jeffrey R. Allen Genetic modification of Gram-negative bacteria to express a desired protein within the cellapos;s periplasmic space, located between the inner cytoplasmic membrane and the outer cell wall, can offer an attractive strategy for commercial production of therapeutic proteins and industrial enzymes. In certain applications, the product expression rate is high, and the ability to isolate the product from the cell mass is greatly enhanced because of the productapos;s compartmentalized location within the cell. Protein release methods that increase the permeability of the outer cell wall for primary recovery, but avoid rupturing the inner cell membrane, reduce contamination of the recovered product with other host cell components and simplify final purification. This article reports representative data for a new release method employing glycol ether solvents. The example involves the use of 2-butoxyethanol (commonly called ethylene glycol n -butyl ether or EB) for selective release of a proprietary biopharmaceutical protein produced in the periplasmic space of Pseudomonas fluorescens. In this example, glycol ether treatment yielded ,65% primary recovery with ,80% purity on a protein-only basis. Compared with other methods including heat treatment, osmotic shock, and the use of surfactants, the glycol ether treatment yielded significantly reduced concentrations of other host cell proteins, lipopolysaccharide endotoxin, and DNA in the recovered protein solution. The use of glycol ethers also allowed exploitation of temperature-change-induced phase splitting behavior to concentrate the desired product. Heating the aqueous EB extract solution to 55 °C formed two liquid phases: a glycol ether-rich phase and an aqueous product phase containing the great majority of the product protein. This phase-splitting step yielded an approximate 10-fold reduction in the volume of the initial product solution and a corresponding increase in the productapos;s concentration. [source] Recent Progress in Biomolecular EngineeringBIOTECHNOLOGY PROGRESS, Issue 1 2000Dewey D. Y. Ryu During the next decade or so, there will be significant and impressive advances in biomolecular engineering, especially in our understanding of the biological roles of various biomolecules inside the cell. The advances in high throughput screening technology for discovery of target molecules and the accumulation of functional genomics and proteomics data at accelerating rates will enable us to design and discover novel biomolecules and proteins on a rational basis in diverse areas of pharmaceutical, agricultural, industrial, and environmental applications. As an applied molecular evolution technology, DNA shuffling will play a key role in biomolecular engineering. In contrast to the point mutation techniques, DNA shuffling exchanges large functional domains of sequences to search for the best candidate molecule, thus mimicking and accelerating the process of sexual recombination in the evolution of life. The phage-display system of combinatorial peptide libraries will be extensively exploited to design and create many novel proteins, as a result of the relative ease of screening and identifying desirable proteins. Even though this system has so far been employed mainly in screening the combinatorial antibody libraries, its application will be extended further into the science of protein-receptor or protein-ligand interactions. The bioinformatics for genome and proteome analyses will contribute substantially toward ever more accelerated advances in the pharmaceutical industry. Biomolecular engineering will no doubt become one of the most important scientific disciplines, because it will enable systematic and comprehensive analyses of gene expression patterns in both normal and diseased cells, as well as the discovery of many new high-value molecules. When the functional genomics database, EST and SAGE techniques, microarray technique, and proteome analysis by 2-dimensional gel electrophoresis or capillary electrophoresis in combination with mass spectrometer are all put to good use, biomolecular engineering research will yield new drug discoveries, improved therapies, and significantly improved or new bioprocess technology. With the advances in biomolecular engineering, the rate of finding new high-value peptides or proteins, including antibodies, vaccines, enzymes, and therapeutic peptides, will continue to accelerate. The targets for the rational design of biomolecules will be broad, diverse, and complex, but many application goals can be achieved through the expansion of knowledge based on biomolecules and their roles and functions in cells and tissues. Some engineered biomolecules, including humanized Mab's, have already entered the clinical trials for therapeutic uses. Early results of the trials and their efficacy are positive and encouraging. Among them, Herceptin, a humanized Mab for breast cancer treatment, became the first drug designed by a biomolecular engineering approach and was approved by the FDA. Soon, new therapeutic drugs and high-value biomolecules will be designed and produced by biomolecular engineering for the treatment or prevention of not-so-easily cured diseases such as cancers, genetic diseases, age-related diseases, and other metabolic diseases. Many more industrial enzymes, which will be engineered to confer desirable properties for the process improvement and manufacturing of high-value biomolecular products at a lower production cost, are also anticipated. New metabolites, including novel antibiotics that are active against resistant strains, will also be produced soon by recombinant organisms having de novo engineered biosynthetic pathway enzyme systems. The biomolecular engineering era is here, and many of benefits will be derived from this field of scientific research for years to come if we are willing to put it to good use. [source] | |||||||||||||