Mesofluidic devices for DNA-programmed combinatorial chemistry

Hybrid combinatorial chemistry strategies that use DNA as an information-carrying medium are proving to be powerful tools for molecular discovery. In order to extend these efforts, we present a highly parallel format for DNA-programmed chemical library synthesis. The new format uses a standard microwell plate footprint and is compatible with commercially available automation technology. It can accommodate a wide variety of combinatorial synthetic schemes with up to 384 different building blocks per chemical step. We demonstrate that fluidic routing of DNA populations in the highly parallel format occurs with excellent specificity, and that chemistry on DNA arrayed into 384 well plates proceeds robustly, two requirements for the high-fidelity translation and efficient in vitro evolution of small molecules.     

Opportunities at the interface of chemistry and biology

The combination of the tools and principles of chemistry, together with the tools of modern molecular biology, allow us to create complex synthetic and natural molecules, and processes with novel biological, chemical and physical properties. This article illustrates the tremendous opportunity that lies at this interface of chemistry and biology by describing a number of examples, ranging from efforts to expand the genetic code of living organisms to the use of combinatorial methods to generate biologically active synthetic molecules.     

Opportunities at the interface of chemistry and biology

The combination of the tools and principles of chemistry, together with the tools of modern molecular biology, allow us to create complex synthetic and natural molecules, and processes with novel biological, chemical and physical properties. This article illustrates the tremendous opportunity that lies at this interface of chemistry and biology by describing a number of examples, ranging from efforts to expand the genetic code of living organisms to the use of combinatorial methods to generate biologically active synthetic molecules.     

Opportunities at the interface of chemistry and biology

The combination of the tools and principles of chemistry, together with the tools of modern molecular biology, allow us to create complex synthetic and natural molecules, and processes with novel biological, chemical and physical properties. This article illustrates the tremendous opportunity that lies at this interface of chemistry and biology by describing a number of examples, ranging from efforts to expand the genetic code of living organisms to the use of combinatorial methods to generate biologically active synthetic molecules.     

Automated synthesis and combinatorial chemistry

The advent of combinatorial chemistry for the high-throughput synthesis of compounds has driven the advancement of new and emerging technologies for synthetic chemistry laboratories. Automated methods for reaction design, information management, chemical synthesis, compound analysis, and biological testing are necessary to realize the full potential of combinatorial chemistry efforts.     

Halogenase engineering and its utility in medicinal chemistry

Halogenation is commonly used in medicinal chemistry to improve the potency of pharmaceutical leads. While synthetic methods for halogenation present selectivity and reactivity challenges, halogenases have evolved over time to perform selective reactions under benign conditions. The optimization of halogenation biocatalysts has utilized enzyme evolution and structure-based engineering alongside biotransformation in a variety of systems to generate stable site-selective variants. The recent improvements in halogenase-catalyzed reactions has demonstrated the utility of these biocatalysts for industrial purposes, and their ability to achieve a broad substrate scope implies a synthetic tractability with increasing relevance in medicinal chemistry.     

Molecular engineering of antimicrobial peptides: microbial targets,peptide motifs and translation opportunities

The global public health threat of antimicrobial resistance has led the scientific community to highly engage into research on alternative strategies to the traditional small molecule therapeutics. Here, we review one of the most popular alternatives amongst basic and applied research scientists, synthetic antimicrobial peptides. The ease of peptide chemical synthesis combined with emerging engineering principles and potent broad-spectrum activity, including against multidrug-resistant strains, has motivated intense scientific focus on these compounds for the past decade. This global effort has resulted in significant advances in our understanding of peptide antimicrobial activity at the molecular scale. Recent evidence of molecular targets other than the microbial lipid membrane, and efforts towards consensus antimicrobial peptide motifs, have supported the rise of molecular engineering approaches and design tools, including machine learning. Beyond molecular concepts, supramolecular chemistry has been lately added to the debate; and helped unravel the impact of peptide self-assembly on activity, including on biofilms and secondary targets, while providing new directions in pharmaceutical formulation through taking advantage of peptide self-assembled nanostructures. We argue that these basic research advances constitute a solid basis for promising industry translation of rationally designed synthetic peptide antimicrobials, not only as novel drugs against multidrug-resistant strains but also as components of emerging antimicrobial biomaterials. This perspective is supported by recent developments of innovative peptide-based and peptide-carrier nanobiomaterials that we also review.     

Exploring the mechanisms of molecular recognition by flavins

The design and development of chemical models for enzymes depends on the fundamental principles observed in biological systems. Molecular recognition of flavins by various receptors has attracted attention due to their applications as chemical models for flavoenzymes. The area of molecular recognition is being investigated through research at the interface of chemistry and biology. In this review, the literature has been surveyed to provide comprehensive coverage of the synthetic methodology of different flavins and receptors and their molecular-recognition studies. Various applications of flavin-receptor complexes have also been highlighted.     

Tools for Selective Enzyme Reaction Steps in the Synthesis of Laboratory Chemicals

The need for more selective reactions steps and the compatibility between process steps which follow on from each other has been a major driving force for organic synthesis. The synthesis of chiral compounds, metabolites, new chemical entities and natural products by a combination of chemical and enzyme reaction steps has become well established, due the existence of stable enzymes as selective catalysts which are inherently chiral by nature. Auxiliary tools such as suitable transfer reagents for reaching complete conversion, easy and robust reaction control as well as tools for straightforward workup and purification of the final product have been developed. Selective enzyme reaction steps in the area of hydrolyses, oxidation steps including hydroxylation and the Baeyer‐Villiger oxidation, carbon‐carbon bond formation and glycosylation reactions have compared favorably with existing methods of classical organic synthesis. The tools developed during optimization and scale‐up of these enzyme reaction steps have the potential to shorten development time. The introduction of selective enzyme reactions into an entire synthetic process has resulted in harmonization of improvements in economic efficiency with resultant solutions to health, safety and environment problems. This will become even more important in industrial synthetic chemistry in the future, for convenient solutions to certain intractable synthetic problems and for expanding the repertoire of chemistry by modular biocatalysts. Efficient isolation procedures for the final product are essential to take full advantage of the biocatalytic conversion to obtain high product yields.     

Synthetic biology: lessons from the history of synthetic organic chemistry

The mid-nineteenth century saw the development of a radical new direction in chemistry: instead of simply analyzing existing molecules, chemists began to synthesize them--including molecules that did not exist in nature. The combination of this new synthetic approach with more traditional analytical approaches revolutionized chemistry, leading to a deep understanding of the fundamental principles of chemical structure and reactivity and to the emergence of the modern pharmaceutical and chemical industries. The history of synthetic chemistry offers a possible roadmap for the development and impact of synthetic biology, a nascent field in which the goal is to build novel biological systems.     

Advances in the chemistry of small molecule fluorescent probes

Small molecule fluorophores are essential tools for chemical biology. A benefit of synthetic dyes is the ability to employ chemical approaches to control the properties and direct the position of the fluorophore. Applying modern synthetic organic chemistry strategies enables efficient tailoring of the chemical structure to obtain probes for specific biological experiments. Chemistry can also be used to activate fluorophores; new fluorogenic enzyme substrates and photoactivatable compounds with improved properties have been prepared that facilitate advanced imaging experiments with low background fluorescence. Finally, chemical reactions in live cells can be used to direct the spatial distribution of the fluorophore, allowing labeling of defined cellular regions with synthetic dyes.     

Towards large-scale synthetic applications of Baeyer-Villiger monooxygenases

Biocatalysis is coming of age, with an increasing number of reactions being scaled-up and developed. The diversity of reactions is also increasing and oxidation reactions have recently been considered for scale-up to commercial processes. One important chemical conversion, which is difficult to achieve enantio- or enantiotopo- selectively, is the Baeyer-Villiger (BV) oxidation of ketones. Using cyclohexanone monooxygenase to catalyse the reaction produces optically pure esters and lactones with exquisite enantiomeric excess values. Recently, these enzymes and their many applications in synthetic chemistry have been explored. The scale-up of these conversions has been examined with the idea of implementing the first commercial Baeyer-Villiger monooxygenase-based process. Here, we review the state-of-the-art situation for the scale-up and exploitation of these enzymes.     

Natural selection in chemical evolution

We propose that chemical evolution can take place by natural selection if a geophysical process is capable of heterotrophic formation of liposomes that grow at some base rate, divide by external agitation, and are subject to stochastic chemical avalanches, in the absence of nucleotides or any monomers capable of modular heredity. We model this process using a simple hill-climbing algorithm, and an artificial chemistry that is unique in exhibiting conservation of mass and energy in an open thermodynamic system. Selection at the liposome level results in the stabilization of rarely occurring molecular autocatalysts that either catalyse or are consumed in reactions that confer liposome level fitness; typically they contribute in parallel to an increasingly conserved intermediary metabolism. Loss of competing autocatalysts can sometimes be adaptive. Steady-state energy flux by the individual increases due to the energetic demands of growth, but also of memory, i.e. maintaining variations in the chemical network. Self-organizing principles such as those proposed by Kauffman, Fontana, and Morowitz have been hypothesized as an ordering principle in chemical evolution, rather than chemical evolution by natural selection. We reject those notions as either logically flawed or at best insufficient in the absence of natural selection. Finally, a finite population model without elitism shows the practical evolutionary constraints for achieving chemical evolution by natural selection in the lab.     

Directed evolution of aldolases for exploitation in synthetic organic chemistry

This review focuses on the directed evolution of aldolases with synthetically useful properties. Directed evolution has been used to address a number of limitations associated with the use of wild-type aldolases as catalysts in synthetic organic chemistry. The generation of aldolase enzymes with a modified or expanded substrate repertoire is described. Particular emphasis is placed on the directed evolution of aldolases with modified stereochemical properties: such enzymes can be useful catalysts in the stereoselective synthesis of biologically active small molecules. The review also describes some of the fundamental insights into mechanistic enzymology that directed evolution can provide.     

Tetrazine ligation for chemical proteomics

Determining small molecule—target protein interaction is essential for the chemical proteomics. One of the most important keys to explore biological system in chemical proteomics field is finding first-class molecular tools. Chemical probes can provide great spatiotemporal control to elucidate biological functions of proteins as well as for interrogating biological pathways. The invention of bioorthogonal chemistry has revolutionized the field of chemical biology by providing superior chemical tools and has been widely used for investigating the dynamics and function of biomolecules in live condition. Among 20 different bioorthogonal reactions, tetrazine ligation has been spotlighted as the most advanced bioorthogonal chemistry because of their extremely faster kinetics and higher specificity than others. Therefore, tetrazine ligation has a tremendous potential to enhance the proteomic research. This review highlights the current status of tetrazine ligation reaction as a molecular tool for the chemical proteomics.     

Uridine natural products: Challenging targets and inspiration for novel small molecule inhibitors

Nucleoside derivatives, in particular those featuring uridine, are familiar components of the nucleoside family of bioactive natural products. The structural complexity and biological activities of these compounds have inspired research from organic chemistry and chemical biology communities seeking to develop novel approaches to assemble the challenging molecular targets, to gain inspiration for enzyme inhibitor development and to fuel antibiotic discovery efforts. This review will present recent case studies describing the total synthesis and biosynthesis of uridine natural products, and de novo synthetic efforts exploiting features of the natural products to produce simplified scaffolds. This research has culminated in the development of complementary strategies that can lead to effective uridine-based inhibitors and antibiotics. The strengths and challenges of the juxtaposing methods will be illustrated by examining select uridine natural products. Moreover, structure–activity relationships (SAR) for each natural product-inspired scaffold will be discussed, highlighting the impact on inhibitor development, with the aim of future uridine-based small molecule expansion.     

Use of model peptide reactions for the characterization of kinetically controlled ligation

Since the introduction of kinetically controlled ligation (KCL), a chemoselective reaction between a peptide-(α)thioarylester and a Cys-peptide-(α)thioalkylester, KCL has been utilized for the total chemical synthesis of large proteins (i.e., lysozyme and HIV-protease) by providing fully convergent synthetic routes. Although KCL has the potential to become an important chemistry for protein synthesis, the principle of KCL is not fully characterized. In particular, prior work on KCL has focused on the reactivity difference of the two different -(α)thioester forms-alkyl vs aryl. Another equally important feature of KCL, Xaa-Cys ligation sites, has not been investigated. The work reported here describes combinatorial KCL reactions using model peptides to dissect the interplay of the Xaa(1), Xaa(2), -(α)thioarylester, and -(α)thioalkylester. Results from these studies provide fundamental insights into the KCL reaction, and will lead to the optimal synthetic route for the routine chemical synthesis of large target protein molecules.     

Exploring the evolution of standard amino-acid alphabet: when genomics meets thermodynamics

One of the most intriguing aspects of life is that despite the diversified apparent shapes, similar building blocks and infrastructures, such as standard amino acids and canonical genetic codes, are shared by most life on Earth. Thus, it is challenging to explore: why nature just selects these building blocks and strategies from numerous candidates to construct life? Was this deterministic or fortuitous? Thanks to the rapid progress in genomics, bioinformatics and synthetic biology, more and more basic principles underlying life design and construction were disclosed in the past decade. However, since the origin of early life is substantially a chemical process, to understand the enigma of life origin, chemists' efforts can not be neglected. In this paper, we focus on the evolution of standard amino-acid alphabet and indicate that chemistry, especially thermodynamics, is indeed critical to understanding the forming mechanisms of amino-acid alphabet. It is revealed that nature prefers low free energy and thus ubiquitous (cheap) small amino acids when beginning to build life, which is compatible with many recent findings from genomics and bioinformatics.     

Prebiotic chemistry in clouds

Summary In the traditional concept for the origin of life as proposed by Oparin and Haldane in the 1920s, prebiotic reactants became slowly concentrated in the primordial oceans and life evolved slowly from a series of highly protracted chemical reactions during the first billion years of Earth's history. However, chemical evolution may not have occurred continuously because planetesimals and asterioids impacted the Earth many times during the first billion years, may have sterilized the Earth, and required the process to start over. A rapid process of chemical evolution may have been required in order that life appeared at or before 3.5 billion years ago. Thus, a setting favoring rapid chemical evolution may be required. A chemical evolution hypothesis set forth by Woese in 1979 accomplished prebiotic reactions rapidly in droplets in giant atmospheric reflux columns. However, in 1985 Scherer raised a number of objections to Woese's hypothesis and concluded that it was not valid. We propose a mechanism for prebiotic chemistry in clouds that satisfies Scherer's concerns regarding the Woese hypothesis and includes advantageous droplet chemistry.Prebiotic reactants were supplied to the atmosphere by comets, meteorites, and interplanetary dust or synthesized in the atmosphere from simple compounds using energy sources such as ultraviolet light, corona discharge, or lightning. These prebiotic monomers would have first encountered moisture in cloud drops and precipitation. We propose that rapid prebiotic chemical evolution was facilitated on the primordial Earth by cycles of condensation and evaporation of cloud drops containing clay condensation nuclei and nonvolatile monomers. For example, amino acids supplied by, or synthesized during entry of, meteorites, comets, and interplanetary dust would have been scavenged by cloud drops containing clay condensation nuclei. Polymerization would have occurred within cloud systems during cycles of condensation, freezing, melting, and evaporation of cloud drops. We suggest that polymerization reactions occurred in the atmosphere as in the Woese hypothesis, but life originated in the ocean as in the Oparin-Haldane hypothesis. The rapidity with which chemical evolution could have occurred within clouds accommodates the time constraints suggested by recent astrophysical theories.