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Multienzymatic system

Zhongwei Yang, Hongling Wang, Yuxiao Wang, Yuhong Ren, Dong-Zhi Wei
The self-assembly of multi-enzyme into bioreactors is of extensive interest to spatially regulate valuable reactions. Despite the important progresses achieved, methods to precisely manufacture multi-enzymatic complex reactors (MECRs) are still poorly proposed both in vivo and in vitro, particularly for more than three bio-catalytically relevant enzymes. Here, we developed a sequential self-assembly system to form multitude MECRs involving three enzymes in the itaconic acid (IA) pathway with two pairs of protein-peptide interactions...
May 4, 2018: ACS Synthetic Biology
Fabián Rico-Rodríguez, Juan Carlos Serrato, Antonia Montilla, Mar Villamiel
Galactooligosaccharides (GOS), recognised prebiotic, can be industrially produced from lactose and commercial β-galactosidase (β-gal) from Kluyveromyces lactis. Residual lactose and glucose limit GOS applications. To handle this problem, a multienzymatic system, with β-gal and glucose oxidase (Gox), was proposed to reduce glucose content in reaction media through its oxidation to gluconic acid (GA). Besides, ultrasound (US) probe effect over the multienzymatic system to produce GOS and GA has been evaluated...
June 2018: Ultrasonics Sonochemistry
Fumiya Iwata, Hidehiko Hirakawa, Teruyuki Nagamune
Bacterial cytochrome P450 monooxygenases (P450s) are promising biocatalysts for chemical syntheses because they catalyze a variety of oxidations on non-activated hydrocarbons using O2 . However, the requirement of two auxiliary proteins, an electron transfer protein and a reductase, for the catalysis is a major bottleneck for in vitro applications of these monooxygenases. Our previous study showed that artificial assembly of a bacterial P450 with its auxiliary proteins using a heterotrimeric proliferating cell nuclear antigen (PCNA) from Sulfolobus solfataricus yields a self-sufficient P450, but partial dissociation of P450 from the complex at catalytic concentrations reduces the apparent specific activity of this self-sufficient P450...
April 16, 2018: Biotechnology Journal
Elizabeth S Lewkowicz, Adolfo M Iribarren
Nucleosides constitute an extensive group of natural and chemically modified compounds that display a wide range of structures and activities. Different biocatalysts have been developed for their preparation, but the choice of commercially available enzymes is limited. Therefore, the search of new biocatalysts is particularly attractive. In this sense, microorganisms are a vast source of enzymatic diversity that can be directly used as a whole cell biocatalysts providing a potential cheaper and suitable route for industrial applications...
October 11, 2017: Current Pharmaceutical Design
Xixian Chen, Congqiang Zhang, Ruiyang Zou, Gregory Stephanopoulos, Heng-Phon Too
In vitro metabolic engineering is an alternative approach to cell-based biosynthesis. It offers unprecedented flexibility for the study of biochemical pathways, thus providing useful information for the rational design and assembly of reaction modules. Herein, we took the advantage of in vitro metabolic engineering to initially gain insight into the regulatory network of a reconstituted amorpha-4,11-diene (AD) synthetic pathway. Guided by lin-log approximation, we rapidly identified the hitherto unrecognized inhibition of adenosine triphosphate (ATP) on both farnesyl pyrophosphate synthase (FPPS) and amorpha-4,11-diene synthase (ADS)...
September 15, 2017: ACS Synthetic Biology
James N Vranish, Mario G Ancona, Eunkeu Oh, Kimihiro Susumu, Igor L Medintz
Enzymes have long been a prime research target for the commercial production of commodity and specialty chemicals, design of sensing devices, and the development of therapeutics and new chemical processes. Industrial applications for enzymes can potentially be enhanced by enzyme immobilization which often allows for increased enzyme stability, facile product purification, and minimized substrate diffusion times in multienzymatic cascades, but this is usually at the cost of a significant decrease in catalytic rates...
April 20, 2017: Nanoscale
Ludmila Martínková, Lenka Rucká, Jan Nešvera, Miroslav Pátek
The aim of this study is to review the current state of and highlight the challenges in the production of microbial nitrilases as catalysts for the mild hydrolysis of industrially important nitriles. Together with aldoxime dehydratase, the nitrile-hydrolyzing enzymes (nitrilase, nitrile hydratase) are key enzymes in the aldoxime-nitrile pathway which is widely distributed in bacteria and fungi. The availability of nitrilases has grown significantly over the past decade due to the use of metagenomic and database-mining approaches...
January 2017: World Journal of Microbiology & Biotechnology
Rosanna Palumbo, Marta Gogliettino, Ennio Cocca, Roberta Iannitti, Annamaria Sandomenico, Menotti Ruvo, Marco Balestrieri, Mosè Rossi, Gianna Palmieri
The proteasome is a multienzymatic complex that controls the half-life of the majority of intracellular proteins, including those involved in apoptosis and cell-cycle progression. Recently, proteasome inhibition has been shown to be an effective anticancer strategy, although its downregulation is often accompanied by severe undesired side effects. We previously reported that the inhibition of acylpeptide hydrolase (APEH) by the peptide SsCEI 4 can significantly affect the proteasome activity in A375 melanoma or Caco-2 adenocarcinoma cell lines, thus shedding new light on therapeutic strategies based on downstream regulation of proteasome functions...
September 23, 2016: International Journal of Molecular Sciences
Frederic Tort, Xènia Ferrer-Cortes, Antonia Ribes
Lipoic acid (LA) is an essential cofactor required for the activity of five multienzymatic complexes that play a central role in the mitochondrial energy metabolism: four 2-oxoacid dehydrogenase complexes [pyruvate dehydrogenase (PDH), branched-chain ketoacid dehydrogenase (BCKDH), 2-ketoglutarate dehydrogenase (2-KGDH), and 2-oxoadipate dehydrogenase (2-OADH)] and the glycine cleavage system (GCS). LA is synthesized in a complex multistep process that requires appropriate function of the mitochondrial fatty acid synthesis (mtFASII) and the biogenesis of iron-sulphur (Fe-S) clusters...
November 2016: Journal of Inherited Metabolic Disease
Julien Diharce, Jérôme Golebiowski, Sébastien Fiorucci, Serge Antonczak
In the course of metabolite formation, some multienzymatic edifices, the so-called metabolon, are formed and lead to a more efficient production of these natural compounds. One of the major features of these enzyme complexes is the facilitation of direct transfer of the metabolite between enzyme active sites by substrate channelling. Biophysical insights into substrate channelling remain scarce because the transient nature of these macromolecular complexes prevents the observation of high resolution structures...
April 21, 2016: Physical Chemistry Chemical Physics: PCCP
Jesús Fernández-Lucas
Living cells are most perfect synthetic factory. The surprising synthetic efficiency of biological systems is allowed by the combination of multiple processes catalyzed by enzymes working sequentially. In this sense, biocatalysis tries to reproduce nature's synthetic strategies to perform the synthesis of different organic compounds using natural catalysts such as cells or enzymes. Nowadays, the use of multienzymatic systems in biocatalysis is becoming a habitual strategy for the synthesis of organic compounds that leads to the realization of complex synthetic schemes...
June 2015: Applied Microbiology and Biotechnology
Leticia S Guidolin, Susana M Morrone Seijo, Francisco F Guaimas, Diego J Comerci, Andrés E Ciocchini
UNLABELLED: Cyclic β-1,2-glucans (CβG) are periplasmic homopolysaccharides that play an important role in the virulence and interaction of Brucella with the host. Once synthesized in the cytoplasm by the CβG synthase (Cgs), CβG are transported to the periplasm by the CβG transporter (Cgt) and succinylated by the CβG modifier enzyme (Cgm). Here, we used a bacterial two-hybrid system and coimmunoprecipitation techniques to study the interaction network between these three integral inner membrane proteins...
May 2015: Journal of Bacteriology
Cheau Yuaan Tan, Hidehiko Hirakawa, Teruyuki Nagamune
Diverse applications of the versatile bacterial cytochrome P450 enzymes (P450s) are hampered by their requirement for the auxiliary proteins, ferredoxin reductases and ferredoxins, that transfer electrons to P450s. Notably, this limits the use of P450s as immobilized enzymes for industrial purposes. Herein, we demonstrate the immobilization of a bacterial P450 and its redox protein partners by supramolecular complex formation using a self-assembled heterotrimeric protein. Employment of homodimeric phosphite dehydrogenase (PTDH) for cross-linking "proliferating cell nuclear antigen-utilized protein complex of P450 and its two electron transfer-related proteins" (PUPPET) yielded a gelling PUPPET-PTDH system capable of regenerating NADH for electron supply owing to its phosphite oxidation activity...
2015: Scientific Reports
Fabio Parmeggiani, Sarah L Lovelock, Nicholas J Weise, Syed T Ahmed, Nicholas J Turner
The synthesis of substituted D-phenylalanines in high yield and excellent optical purity, starting from inexpensive cinnamic acids, has been achieved with a novel one-pot approach by coupling phenylalanine ammonia lyase (PAL) amination with a chemoenzymatic deracemization (based on stereoselective oxidation and nonselective reduction). A simple high-throughput solid-phase screening method has also been developed to identify PALs with higher rates of formation of non-natural D-phenylalanines. The best variants were exploited in the chemoenzymatic cascade, thus increasing the yield and ee value of the D-configured product...
April 7, 2015: Angewandte Chemie
Magdalena Kotowska, Krzysztof Pawlik
A large number of antibiotics and other industrially important microbial secondary metabolites are synthesized by polyketide synthases (PKSs) and nonribosomal peptide synthetases (NRPSs). These multienzymatic complexes provide an enormous flexibility in formation of diverse chemical structures from simple substrates, such as carboxylic acids and amino acids. Modular PKSs and NRPSs, often referred to as megasynthases, have brought about a special interest due to the colinearity between enzymatic domains in the proteins working as an "assembly line" and the chain elongation and modification steps...
September 2014: Applied Microbiology and Biotechnology
Iris Mangas, Eugenio Vilanova, Jorge Estévez
Phenylmethylsulfonyl fluoride (PMSF) is a protease and esterase inhibitor that causes protection, or potentiation/"promotion," of organophosphorus delayed neuropathy (OPIDN), depending on whether it is dosed before or after an inducer of delayed neuropathy, such as mipafox. The molecular target of the potentiation/promotion of OPIDN has not yet been identified. The kinetic data of phenyl valerate esterase inhibition by PMSF were obtained with membrane chicken brain fractions, the animal model and tissue in which neuropathy target esterase (NTE) was first described...
February 2014: Archives of Toxicology
Inés Ardao, Ee Taek Hwang, An-Ping Zeng
: In vitro multienzymatic bioreaction systems are attracting increasing attention for the development of bioproduction systems. The de-coupling of the biocatalytic pathway from the cellular machinery aimed at growth and survival allows achievement of high product yields and thus reduces byproduct or waste generation. Additionally, the use of several enzymes allows the realization of much more complex synthetic schemes, thus expanding the chemical diversity of synthetic compounds with new chemical properties or bioactivities...
2013: Advances in Biochemical Engineering/biotechnology
Milja Pešić, Carmen López, Josep López-Santín, Gregorio Alvaro
In this work, the successful coupling of enzymatic oxidation and aldol addition reactions for the synthesis of a Cbz-aminopolyol from a Cbz-amino alcohol was achieved for the first time in a multienzymatic one-pot system. The two-step cascade reaction consisted of the oxidation of Cbz-ethanolamine to Cbz-glycinal catalyzed by chloroperoxidase from the fungus Caldariomyces fumago and aldol addition of dihydroxyacetone phosphate to Cbz-glycinal catalyzed by rhamnulose-1-phosphate aldolase expressed as a recombinant enzyme in Escherichia coli, yielding (3R,4S)-5-{[(benzyloxy)carbonyl]amino}-5-deoxy-1-O-phosphonopent-2-ulose...
August 2013: Applied Microbiology and Biotechnology
Elisabetta Brenna, Francesco G Gatti, Luciana Malpezzi, Daniela Monti, Fabio Parmeggiani, Alessandro Sacchetti
A stereoselective synthesis of bicyclic primary or secondary amines, based on tetralin or chroman structural moieties, is reported. These amines are precursors of important active pharmaceutical ingredients such as rotigotine (Neupro), robalzotan, and ebalzotan. The key step is based on a multienzymatic reduction of an α,β-unsaturated aldehyde or ketone to give the saturated primary or secondary alcohol, in a high yield and with a high ee. The catalytic system consists of the combination of an ene-reductase (ER; i...
May 17, 2013: Journal of Organic Chemistry
Ildefonso M De la Fuente, Jesus M Cortes, David A Pelta, Juan Veguillas
BACKGROUND: The experimental observations and numerical studies with dissipative metabolic networks have shown that cellular enzymatic activity self-organizes spontaneously leading to the emergence of a Systemic Metabolic Structure in the cell, characterized by a set of different enzymatic reactions always locked into active states (metabolic core) while the rest of the catalytic processes are only intermittently active. This global metabolic structure was verified for Escherichia coli, Helicobacter pylori and Saccharomyces cerevisiae, and it seems to be a common key feature to all cellular organisms...
2013: PloS One
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