scholarly journals Genetically Encodable Scaffolds for Optimizing Enzyme Function

Molecules ◽  
2021 ◽  
Vol 26 (5) ◽  
pp. 1389
Author(s):  
Yong Quan Tan ◽  
Bo Xue ◽  
Wen Shan Yew
Keyword(s):  
Synthetic Biology ◽  
Metabolic Flux ◽  
Enzyme Stability ◽  
Enzyme Engineering ◽  
Enzyme Function ◽  
Enzyme Properties ◽  
Molecular Tools ◽  
Reaction Conditions ◽  
Recent Trends ◽  
Indispensable Tool

Enzyme engineering is an indispensable tool in the field of synthetic biology, where enzymes are challenged to carry out novel or improved functions. Achieving these goals sometimes goes beyond modifying the primary sequence of the enzyme itself. The use of protein or nucleic acid scaffolds to enhance enzyme properties has been reported for applications such as microbial production of chemicals, biosensor development and bioremediation. Key advantages of using these assemblies include optimizing reaction conditions, improving metabolic flux and increasing enzyme stability. This review summarizes recent trends in utilizing genetically encodable scaffolds, developed in line with synthetic biology methodologies, to complement the purposeful deployment of enzymes. Current molecular tools for constructing these synthetic enzyme-scaffold systems are also highlighted.

scholarly journals Application of enzyme engineering in pharmacy

2020 ◽  
Vol 9 (2) ◽  
Author(s):  
Qiaole Li ◽  
Yanhong Li ◽  
Haoran Niu
Keyword(s):  
High Efficiency ◽  
Development Trend ◽  
Enzyme Engineering ◽  
Reaction Conditions ◽  
Important Method ◽  
Reaction Specificity ◽  
Prevention And Treatment ◽  
Modern Biotechnology ◽  
Catalytic Functions ◽  
The Development Trend

Enzyme engineering is an important part of modern biotechnology. Due to its high reaction specificity, high efficiency, mild reaction conditions, and low pollution, it is also an important method widely used in the pharmaceutical field. The application of enzymes in medicine is diverse, such as: diagnosis, prevention and treatment of diseases with enzymes, manufacture of various drugs with enzymes, etc., mainly through manual operations, to obtain enzymes required by the pharmaceutical industry, and through various means Enzymes perform their catalytic functions. This article mainly introduces the application of enzyme engineering in the pharmaceutical field, and also prospects the development trend of enzyme engineering in the pharmaceutical field.

Cell-free synthetic biology in the new era of enzyme engineering

2020 ◽  
Vol 28 (11) ◽  
pp. 2810-2816
Author(s):  
Nan Jiang ◽  
Lianju Ma ◽  
Yuan Lu
Keyword(s):  
Synthetic Biology ◽  
Enzyme Engineering ◽  
New Era

scholarly journals Engineering polymerases for applications in synthetic biology

2020 ◽  
Vol 53 ◽  
Author(s):  
Ali Nikoomanzar ◽  
Nicholas Chim ◽  
Eric J. Yik ◽  
John C. Chaput
Keyword(s):  
Cell Division ◽  
Synthetic Biology ◽  
Substrate Specificity ◽  
Physicochemical Properties ◽  
Genetic Information ◽  
Biomedical Applications ◽  
Dna Polymerases ◽  
Enzyme Engineering ◽  
Practical Applications ◽  
Naturally Occurring

Abstract DNA polymerases play a central role in biology by transferring genetic information from one generation to the next during cell division. Harnessing the power of these enzymes in the laboratory has fueled an increase in biomedical applications that involve the synthesis, amplification, and sequencing of DNA. However, the high substrate specificity exhibited by most naturally occurring DNA polymerases often precludes their use in practical applications that require modified substrates. Moving beyond natural genetic polymers requires sophisticated enzyme-engineering technologies that can be used to direct the evolution of engineered polymerases that function with tailor-made activities. Such efforts are expected to uniquely drive emerging applications in synthetic biology by enabling the synthesis, replication, and evolution of synthetic genetic polymers with new physicochemical properties.

scholarly journals Enzymatic Bioreactors: An Electrochemical Perspective

Catalysts ◽  
2020 ◽  
Vol 10 (11) ◽  
pp. 1232
Author(s):  
Simin Arshi ◽  
Mehran Nozari-Asbemarz ◽  
Edmond Magner
Keyword(s):  
Direct Electron Transfer ◽  
High Surface ◽  
High Surface Area ◽  
Enzyme Engineering ◽  
Future Perspective ◽  
Enzymatic System ◽  
Reaction Conditions ◽  
Flow Reactors ◽  
Recent Developments ◽  
Enzyme Cascades

Biocatalysts provide a number of advantages such as high selectivity, the ability to operate under mild reaction conditions and availability from renewable resources that are of interest in the development of bioreactors for applications in the pharmaceutical and other sectors. The use of oxidoreductases in biocatalytic reactors is primarily focused on the use of NAD(P)-dependent enzymes, with the recycling of the cofactor occurring via an additional enzymatic system. The use of electrochemically based systems has been limited. This review focuses on the development of electrochemically based biocatalytic reactors. The mechanisms of mediated and direct electron transfer together with methods of immobilising enzymes are briefly reviewed. The use of electrochemically based batch and flow reactors is reviewed in detail with a focus on recent developments in the use of high surface area electrodes, enzyme engineering and enzyme cascades. A future perspective on electrochemically based bioreactors is presented.

Preparation of aqueous polyaniline-vesicle suspensions with class III peroxidases. Comparison between horseradish peroxidase isoenzyme C and soybean peroxidase

2013 ◽  
Vol 67 (8) ◽  
Author(s):  
Katja Junker ◽  
Ivan Gitsov ◽  
Nick Quade ◽  
Peter Walde
Keyword(s):  
Horseradish Peroxidase ◽  
Enzyme Stability ◽  
Class Iii ◽  
Salt Form ◽  
Reaction Conditions ◽  
Soybean Peroxidase ◽  
Peroxidase Isoenzyme ◽  
Complete Inactivation ◽  
Class Iii Peroxidases ◽  
Optimal Reaction

AbstractAniline was polymerised enzymatically in aqueous solution at pH = 4.3 and 25°C in the presence of submicrometer-sized vesicles formed from sodium bis(2-ethylhexyl)sulphosuccinate (AOT). H2O2 served as oxidant and the enzyme used was either horseradish peroxidase isoenzyme C (HRPC) or soybean peroxidase (SBP), both being class III peroxidases. From previous studies with HRPC, it is known that stable vesicle suspensions containing the emeraldine salt form of polyaniline (PANI-ES) can be obtained within 1–2 days with a 90–95 % yield, provided that optimal reaction conditions are applied. Unfortunately, HRPC becomes inactivated during polymerisation. In the present study, a linear dendritic block copolymer was added to HRPC, resulting in higher operational enzyme stability; the stabilising effect, however, was too small to afford a substantial decrease in the required amount of enzyme. Moreover, replacing HRPC with SBP was of no advantage, although SBP is known to be more stable towards inactivation by H2O2 than HRPC. By contrast, SBP was found to be much slower in oxidising aniline, and complete inactivation of SBP occurred before all the aniline monomers were oxidised, leading to low yields and the formation of over-oxidised products. The same was observed for HRP isoenzyme A2. Reactions without vesicles indicated that peroxidase inactivation was probably caused by PANI-ES.

scholarly journals Recent Trends and Applications of Molecular Modeling in GPCR–Ligand Recognition and Structure-Based Drug Design

2018 ◽  
Vol 19 (7) ◽  
pp. 2105 ◽  
Author(s):  
Xiaojing Yuan ◽  
Yechun Xu
Keyword(s):  
Molecular Modeling ◽  
Drug Design ◽  
Ligand Recognition ◽  
Energy Calculation ◽  
Enhanced Sampling ◽  
Structure Based Drug Design ◽  
Recent Trends ◽  
Indispensable Tool ◽  
G Protein Coupled

G protein-coupled receptors represent the largest family of human membrane proteins and are modulated by a variety of drugs and endogenous ligands. Molecular modeling techniques, especially enhanced sampling methods, have provided significant insight into the mechanism of GPCR–ligand recognition. Notably, the crucial role of the membrane in the ligand-receptor association process has earned much attention. Additionally, docking, together with more accurate free energy calculation methods, is playing an important role in the design of novel compounds targeting GPCRs. Here, we summarize the recent progress in the computational studies focusing on the above issues. In the future, with continuous improvement in both computational hardware and algorithms, molecular modeling would serve as an indispensable tool in a wider scope of the research concerning GPCR–ligand recognition as well as drug design targeting GPCRs.

scholarly journals A Streptomyces venezuelae Cell-Free Toolkit for Synthetic Biology

2020 ◽  
Author(s):  
Simon J Moore ◽  
Hung-En Lai ◽  
Soo-Mei Chee ◽  
Ming Toh ◽  
Seth Coode ◽  
...  
Keyword(s):  
Synthetic Biology ◽  
Gene Clusters ◽  
Test Tube ◽  
High Yield ◽  
Proof Of Concept ◽  
Biosynthetic Gene Clusters ◽  
Reaction Conditions ◽  
One Pot ◽  
Free System ◽  
High Level

AbstractProkaryotic cell-free coupled transcription-translation (TX-TL) systems are emerging as a powerful tool to examine natural product biosynthetic pathways in a test-tube. The key advantages of this approach are the reduced experimental timescales and controlled reaction conditions. In order to realise this potential, specialised cell-free systems in organisms enriched for biosynthetic gene clusters, with strong protein production and well-characterised synthetic biology tools, is essential. The Streptomyces genus is a major source of natural products. To study enzymes and pathways from Streptomyces, we originally developed a homologous Streptomyces cell-free system to provide a native protein folding environment, a high G+C (%) tRNA pool and an active background metabolism. However, our initial yields were low (36 μg/mL) and showed a high level of batch-to-batch variation. Here, we present an updated high-yield and robust Streptomyces TX-TL protocol, reaching up to yields of 266 μg/mL of expressed recombinant protein. To complement this, we rapidly characterise a range of DNA parts with different reporters, express high G+C (%) biosynthetic genes and demonstrate an initial proof of concept for combined transcription, translation and biosynthesis of Streptomyces metabolic pathways in a single ‘one-pot’ reaction.

scholarly journals Towards developing algal synthetic biology

2016 ◽  
Vol 44 (3) ◽  
pp. 716-722 ◽  
Author(s):  
Mark Aden Scaife ◽  
Alison Gail Smith
Keyword(s):  
Synthetic Biology ◽  
Transgene Expression ◽  
Phaeodactylum Tricornutum ◽  
Genetic Manipulation ◽  
Lessons Learned ◽  
Molecular Tools ◽  
Metabolic Diversity ◽  
Design Build ◽  
Fundamental Research ◽  
And Function

The genetic, physiological and metabolic diversity of microalgae has driven fundamental research into photosynthesis, flagella structure and function, and eukaryotic evolution. Within the last 10 years these organisms have also been investigated as potential biotechnology platforms, for example to produce high value compounds such as long chain polyunsaturated fatty acids, pigments and antioxidants, and for biodiesel precursors, in particular triacylglycerols (TAGs). Transformation protocols, molecular tools and genome sequences are available for a number of model species including the green alga Chlamydomonas reinhardtii and the diatom Phaeodactylum tricornutum, although for both species there are bottlenecks to be overcome to allow rapid and predictable genetic manipulation. One approach to do this would be to apply the principles of synthetic biology to microalgae, namely the cycle of Design-Build-Test, which requires more robust, predictable and high throughput methods. In this mini-review we highlight recent progress in the areas of improving transgene expression, genome editing, identification and design of standard genetic elements (parts), and the use of microfluidics to increase throughput. We suggest that combining these approaches will provide the means to establish algal synthetic biology, and that application of standard parts and workflows will avoid parallel development and capitalize on lessons learned from other systems.

scholarly journals Active-site loop variations adjust activity and selectivity of the cumene dioxygenase

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Peter M. Heinemann ◽  
Daniel Armbruster ◽  
Bernhard Hauer
Keyword(s):  
Active Site ◽  
Hot Spots ◽  
Degrees Of Freedom ◽  
Enzyme Engineering ◽  
Enzyme Properties ◽  
Sequence Alignments ◽  
Alanine Scan ◽  
High Flexibility ◽  
Important Enzyme ◽  
Novel Products

AbstractActive-site loops play essential roles in various catalytically important enzyme properties like activity, selectivity, and substrate scope. However, their high flexibility and diversity makes them challenging to incorporate into rational enzyme engineering strategies. Here, we report the engineering of hot-spots in loops of the cumene dioxygenase from Pseudomonas fluorescens IP01 with high impact on activity, regio- and enantioselectivity. Libraries based on alanine scan, sequence alignments, and deletions along with a novel insertion approach result in up to 16-fold increases in activity and the formation of novel products and enantiomers. CAVER analysis suggests possible increases in the active pocket volume and formation of new active-site tunnels, suggesting additional degrees of freedom of the substrate in the pocket. The combination of identified hot-spots with the Linker In Loop Insertion approach proves to be a valuable addition to future loop engineering approaches for enhanced biocatalysts.

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