Acid Side Chains Research Articles

Divergent Metabolism of Cyclo-l-Trp-l-Leu in Streptomyces albofaciens by Hydroxylation and Nucleobase Transfer with Two Cytochrome P450 Enzymes.

A three-gene salb cluster from Streptomyces albofaciens was proven to be responsible for the formation of cyclo-l-Trp-l-Leu (cWL) derivatives. An Escherichia coli strain harboring the cyclodipeptide synthase (CDPS) gene salbA produced cWL. Expression of the whole cluster or genes of various combinations in Streptomyces coelicolor revealed different metabolites of cWL by two cytochrome P450 enzymes. Isolation and structure elucidation proved the conversion of cWL to guatrypleumycine A by nucleobase transfer with SalbB and to cyclo(trans-10-hydroxy-l-Trp-l-Leu) by hydroxylation with SalbC. Incubation with 15NH4Cl supported the incorporation of guanine in guatrypleumycine A and an X-ray crystallographic study confirmed the stereospecific hydroxylation at C-10 of the tryptophanyl residue. Cultivation of the salbB or salbC expression strains with different substrates further proved the divergent metabolisms of cWL. To the best of our knowledge, SalbC is the first report of the P450 enzyme from CDPS-associated pathways to catalyze β-hydroxylation at the amino acid side chain.

Substitution of the Axial Type 1 Cu Ligand Affords Binding of a Water Molecule in Axial Position Affecting Kinetics, Spectral, and Structural Properties of the Small Laccase Ssl1.

Multicopper oxidases use Cu ions as cofactors to oxidize various substrates. High reduction potential at Type 1 Cu is considered as crucial for effective catalysis. Previous studies have shown that replacing the axial methionine ligand of the Type 1 Cu with leucine or phenylalanine leads to an increased reduction potential, but not always to higher enzyme activity. Here we present a study on six variants of the small laccase Ssl1 from Streptomyces sviceus, where the axial methionine ligand was substituted, and the effect of the axial ligand on reduction potential, activity, spectral properties and structure was investigated. Absorption, electronic circular dichroism and EPR spectra revealed the presence of a stronger coordinating axial ligand like oxygen, which influences the electronic and catalytic properties more than the nature of the amino acid side chain. The crystal structures of the Ssl1 variants were solved, which show that none of the amino acid side chains coordinate to the Cu. Instead, a water molecule is found in the axial coordination site, which support the spectroscopic data. Our findings highlight the importance of combining structural and spectroscopic methods to investigate the effect of amino acid exchange on multicopper oxidases.

Controlled Helical Organization in Supramolecular Polymers of Pseudo-Macrocyclic Tetrakisporphyrins.

Tetrakisporphyrin monomers with amino acid side chains at each end form intramolecular antiparallel hydrogen-bonds to adopt chirally twisted pseudo-macrocyclic structures that result in right-handed and left-handed (P)- and (M)-conformations. The pseudo-macrocyclic tetrakisporphyrin monomers self-assembled to form supramolecular helical pseudo-polycatenane polymers via head-to-head complementary dimerization of the bisporphyrin cleft units in an isodesmic manner. The formation of one-handed supramolecular helical pseudo-polycatenane polymers was confirmed by circular dichroism (CD) spectroscopy. The methyl and iso-propyl groups at the stereogenic center greatly enhanced the induced circular dichroism in the Soret bands of the supramolecular helical pseudo-polycatenane polymers. The induced CDs were reduced upon the introduction of large iso-butyl and tert-butyl groups. Atomic force microscopy revealed well-grown and long supramolecular helical pseudo-polycatenane polymer chains with chain lengths in the range of 361 to 13.6 nm. The right-handed helical chains were established by the self-assembly of the right-handed (P)-conformation of the pseudo-macrocyclic monomer.

Controlled Helical Organization in Supramolecular Polymers of Pseudo‐Macrocyclic Tetrakisporphyrins

AbstractTetrakisporphyrin monomers with amino acid side chains at each end form intramolecular antiparallel hydrogen‐bonds to adopt chirally twisted pseudo‐macrocyclic structures that result in right‐handed and left‐handed (P)‐ and (M)‐conformations. The pseudo‐macrocyclic tetrakisporphyrin monomers self‐assembled to form supramolecular helical pseudo‐polycatenane polymers via head‐to‐head complementary dimerization of the bisporphyrin cleft units in an isodesmic manner. The formation of one‐handed supramolecular helical pseudo‐polycatenane polymers was confirmed by circular dichroism (CD) spectroscopy. The methyl and iso‐propyl groups at the stereogenic center greatly enhanced the induced circular dichroism in the Soret bands of the supramolecular helical pseudo‐polycatenane polymers. The induced CDs were reduced upon the introduction of large iso‐butyl and tert‐butyl groups. Atomic force microscopy revealed well‐grown and long supramolecular helical pseudo‐polycatenane polymer chains with chain lengths in the range of 361 to 13.6 nm. The right‐handed helical chains were established by the self‐assembly of the right‐handed (P)‐conformation of the pseudo‐macrocyclic monomer.

Likely Overstabilization of Charge-Charge Interactions in CHARMM36m(w): A Case for a99SB-disp Water.

Recent years have witnessed drastic improvements in general-purpose explicit solvent protein force fields, partially driven by the need to study intrinsically disordered proteins (IDPs), and yet the state-of-the-art force fields such as CHARMM36m (c36m) and a99SB-disp still provide different performances in simulating disordered protein states, where c36m has a bias toward overcompaction for large IDPs. Here, we examine the performance of c36m and a99SB-disp in describing the stabilities of a set of 46 amino acid backbone and side chain pairs in various configurations. The free energy results show that c36m systematically predicts stronger interactions compared to a99SB-disp by an average of 0.2 kcal/mol for nonpolar pairs, 0.6 kcal/mol for polar pairs, and 0.8 kcal/mol for salt bridges. The most severe overstabilization in c36m is observed for charged pairs involving the Arg and Glu side chains by up to 2.9 kcal/mol. Importantly, the systematic overstabilization of c36m is only marginally alleviated by c36mw, an ad hoc patch to c36m that increases the dispersion interactions between TIP3P hydrogens and protein atoms. Guided by free energy decomposition, we evaluated if revising the charges alone could alleviate the severe overstabilization of salt bridges of c36m(w) vs a99SB-disp. The results suggested that the direct modification of protein-water interactions is also necessary. Toward this end, we proposed a tentative modification to c36m, referred to as c36mrb-disp, which combines modified Arg side chain charges, retuned backbone hydrogen bonding strength, and the a99SB-disp water model. The modified force field successfully reproduces the secondary structures of several intrinsically disordered peptides and proteins including (AAQAA)3, GB1p, and p53 transactivation domain, while maintaining the stability of a set of folded proteins. This work provides a set of useful systems for benchmarking and optimizing protein force fields and highlights the importance of balancing protein-protein and protein-water electrostatic interactions for accurately describing both folded and disordered proteins.

Long-Range Proton Channels Constructed via Hierarchical Peptide Self-Assembly.

The quest to understand and mimic proton translocation mechanisms in natural channels has driven the development of peptide-based artificial channels facilitating efficient proton transport across nanometric membranes. It is demonstrated here that hierarchical peptide self-assembly can form micrometers-long proton nanochannels. The fourfold symmetrical peptide design leverages intermolecular aromatic interactions to align self-assembled cyclic peptide nanotubes, creating hydrophilic nanochannels between them. Titratable amino acid sidechains are positioned adjacent to each other within the channels, enabling the formation of hydrogen-bonded chains upon hydration, and facilitating efficient proton transport. Moreover, these chains are enriched with protons and water molecules by interacting with immobile counter ions introduced into the channels, increasing proton flow density and rate. This system maintains proton transfer rates closely resembling those in natural protein channels over micrometer distances. The functional behavior of these inherently recyclable and biocompatible systems opens the door for their exploitation in diverse applications in energy storage and conversion, biomedicine, and bioelectronics.

A High-Performance Polyimide Composite Membrane with Flexible Polyphosphazene Derivatives for Vanadium Redox Flow Battery.

A sulfonated polyimide, S-F-abSPI, with alkyl sulfonic acid side chains, and a polyphosphonitrile derivative, poly[4-methoxyphenoxy (4-fluorophenoxy) phosphazene] (PFMPP), were designed and synthesized. Composite modification of the S-F-abSPI membrane was carried out using PFMPP, resulting in the preparation of composite membranes with different composite ratios, which were then subjected to performance testing and characterization. Experimental results revealed a significant enhancement in the proton conductivity of the S-F-abSPI membrane, reaching 0.116 S cm-1, slightly higher than that of the N212 membrane. The S-F-abSPI/1% PFMPP composite membrane exhibited the optimal comprehensive performance, with a surface resistance as low as 0.54 Ω cm2, comparable to that of the N212 membrane. At a high current density of 200 mA cm-2 during charge-discharge, the composite membrane achieved a voltage efficiency (VE) of 83.12% and an energy efficiency (EE) of 81.95%. Cycling tests over 200 cycles demonstrated the composite membrane's excellent long-term cycling stability. The alkyl sulfonic acid side chains enhanced the proton conductivity of the membrane, while electrostatic potential distribution calculations indicated strong interactions between PFMPP and the base membrane, enhancing the membrane's mechanical strength, reducing vanadium ion permeability, and improving chemical stability and vanadium ion selectivity. This composite membrane holds promise for high-performance VRFB applications.

Feather keratin in Pavo cristatus: A tentative structure

Abstract Background F-keratin forms an evolutionary conserved nanocomposite which allows birds to fly. Structural models for F-keratin are all based on the pioneering X-ray diffraction studies on seagull F-keratin, first published in 1932, confirmed for other species and refined over the years. There is, however, no experimental proof because native F-keratin does not form a perfect molecular crystal as required for structure determination. Methods Peacock’s tail feathers were systematically re-investigated by taking diffraction patterns at different rotation angles. Using the recently developed AlphaFold algorithm, a collection of 3D models of arbitrarily truncated and multiplied Pavo cristatus F-keratin sequences was created. The shape, dimensions, density and interfacial exposure of functionally relevant amino acid side chains of the calculated 3D building blocks were used as the initial selection criteria for filamentous F-keratin precursors. Full reproducibility of in silico folding and agreement with previous results from mechanical testing, biochemical analyses and SAXS experiments was mandatory for suggesting the tentative structure for the novel F-keratin repeating unit. Results The filament of the F-keratin polymer is an alternating arrangement of two units called "N-block" and "C-block": Four strands AA 1–52 form a disulfide-stabilized twisted parallelepiped, with 89° internal rotation within eight levels of β-sandwiches. Four strands AA 81–100 form a two-level “β-sandwich” in which aromatic residues provide resilience, like vertebral discs in a spinal column. The pitch of an N+C-block octamer is 10 nm. Solidification may involve "C-blocks" to temporarily mold into "C-wedges" of 18° tilt, which align F-keratin into laterally amorphous fiber-reinforced composites of 9.5 nm axial periodicity. This experimentally significant distance corresponds to the fully stretched AA 53–80 matrix segment. The deformed “spinal column” unwinds under compression when F-keratin filaments perfectly align horizontally into stacked sheets in the solid state. Conclusions At present, the tentative structure presented here is without alternatives.

Light-Activated Reactivity of Nitrones with Amino Acids and Proteins.

Controlled modifications of amino acids are an indispensable tool for advancing fundamental and translational research based on peptides and proteins. Yet, we still lack methods to chemically modify each naturally occurring amino acid sidechain. To help address this gap, we show that N,α-diaryl oxaziridines expand the scope of bioconjugation methods to chemically modify cysteine, methionine, and tryptophan residues with evidence for additional tyrosine labelling in a proteomic context. Conjugation primarily at tryptophan sites can be accessed by selective cleavage of modifications at other sidechains. The N,α-diaryl oxaziridine reagents are accessed through photoisomerization of nitrones, which serve as photocaged reagents, thus providing an additional level of control over reactivity. Initial guiding principles for the design of nitrone reagents are developed by exploring the impact of structure on UV-vis absorption, photoisomerization, and reactivity. We identify a nitrone structure that maximizes photoisomerization efficiency, the aqueous stability of the oxaziridine, the extent of amino acid modification, and the stability of the resulting amino acid conjugates. We then translate nitrone reagents to modify proteins in aqueous conditions. Finally, we use nitrones to profile reactive residues across the proteome of a mammalian cell line and find that they expand the proteome coverage.

Peptide Aldehydes Incorporating Thiazol-4-yl Alanine Are Potent In Vitro Inhibitors of SARS-CoV-2 Main Protease.

The main protease of SARS-CoV-2 is an essential enzyme required for polyprotein cleavage during viral replication and thus is an excellent target for development of direct-acting antiviral compounds. Continued research efforts have elucidated several peptidic small molecules like GC376, boceprevir, and nirmatrelvir with potent anticoronaviral activity bearing optimized amino acid side chain residues. To reduce synthetic complexity and cost, we used simple chemical surrogates that were commercially readily available to develop new inhibitors that mimic the potency of these drug compounds. We synthesized and tested several analogue chimeras of GC376 and boceprevir that have surrogate residues at the P1 and/or P2 position in order to further improve target binding. Both P1 variants with either a nonpolar cyclobutyl or polar thiazol-4-yl alanine resulted in low-micromolar to submicromolar Mpro inhibitors with strong antiviral activity in cell assays.

Efficient generation of protein pockets with PocketGen

Designing protein-binding proteins is critical for drug discovery. However, artificial-intelligence-based design of such proteins is challenging due to the complexity of protein–ligand interactions, the flexibility of ligand molecules and amino acid side chains, and sequence–structure dependencies. We introduce PocketGen, a deep generative model that produces residue sequence and atomic structure of the protein regions in which ligand interactions occur. PocketGen promotes consistency between protein sequence and structure by using a graph transformer for structural encoding and a sequence refinement module based on a protein language model. The graph transformer captures interactions at multiple scales, including atom, residue and ligand levels. For sequence refinement, PocketGen integrates a structural adapter into the protein language model, ensuring that structure-based predictions align with sequence-based predictions. PocketGen can generate high-fidelity protein pockets with enhanced binding affinity and structural validity. It operates ten times faster than physics-based methods and achieves a 97% success rate, defined as the percentage of generated pockets with higher binding affinity than reference pockets. Additionally, it attains an amino acid recovery rate exceeding 63%.

The Molecular Design of a Macrocycle Descaling Agent Based on Azacrown and the Mechanism of Barium Sulfate Scale Removal.

The formation of barium sulfate scale is a persistent and formidable challenge across various industrial processes. In order to effectively mitigate this problem, this study proposed the development of an innovative azacrown ether-based macrocycle descaling agent. Using density functional theory, an in-depth analysis of the surface energy of different barium sulfate crystal facets was carried out, together with a detailed investigation into the adsorption properties of the functional groups on the (001) surface. A further comprehensive investigation was carried out to determine how changes in the nitrogen and oxygen atoms in the crown ether framework influence its adsorption affinity to barium ions. In addition, a detailed analysis was carried out to elucidate the molecular interactions between crown ethers with pyridine carboxylic acid side chains and barium sulfate. The newly developed decalcifying macrocycle descaling agent exhibited superior adsorption performance, achieving an adsorption energy for barium ions approximately -4.1512 ev higher than that of conventional DTPA decalcifiers. This remarkable improvement is mainly attributed to the pivotal role of electrostatic forces in the coordination process between the macrocycle descaling agent and barium ions, with an electrostatic potential value reaching -143.37 kcal/mol. This discovery not only introduces a novel approach to the removal of barium sulfate scale but also highlights the significant potential of macrocycle chemistry in industrial applications.

Nucleotide-mediated modulation of chemoselective protein functionalization in a liquid-like condensed phase

Liquid-like protein condensates are ubiquitous in cellular system and are increasingly recognized for their roles in physiological processes. Condensed phase harbors distinctive chemical microenvironment, markedly different than dilute aqueous phase. Herein, we demonstrate chemoselective modification pattern of nucleophilic canonical amino acid sidechains (namely – cysteine, tyrosine and lysine) of the protein towards 4-chloro-7-nitrobenzofurazan in the dilute and condensed phase. We also delineate how the effect of nucleotides and their in situ enzymatic dissociation temporally modulate the protein condensate’s pH and the protein’s corresponding chemoselective modification. We have shown that the pH of the condensate decreases in the presence of nucleoside triphosphate, whereas it increases in the presence of nucleoside monophosphates or phosphate ion. For instance, we find lysine-specific modification gets inhibited in the presence of adenosine triphosphate (ATP), but significantly enhanced in the presence of monophosphates. This feature enables us to gain temporal control over dynamic change in protein functionalization via enzymatic ATP hydrolysis. Overall, this work substantiates the alteration in pH-responsiveness of Brønsted basicity of a protein’s ε-amine in the condensed phase. Furthermore, this environment sensitivity in chemoselective protein functionalization in condensed phase will be important in adaptable protein engineering to the chemical biology of protein phase separation.

Efficiency Enhancement in Peptide Hydrogelators: The Crucial Role of Side Chain Hydrogen Bonding Over Aromatic π-π Interactions.

Short peptide assemblies that form supramolecular hydrogels are stabilized by both intermolecular noncovalent interactions among amino acid side chains and hydrogen bonding between peptide backbone amides. Previous research has emphasized the inclusion of aromatic amino acids in short peptide sequences, positing that aromatic π-π interactions contribute significantly to inducing efficient hydrogelation i.e., at low minimum gelation concentrations (MGCs). However, herein, we demonstrate that additional hydrogen bonding interactions from amino acid side chains play a more pivotal role in the efficiency of peptide hydrogelation compared to aromatic π-π interactions. We investigated two sets of Fluorenylmethoxycarbonyl (Fmoc)-functionalized α-synuclein and human islet amyloid peptide fragments [Fmoc-NVGGAVVT (Syn-N) and Fmoc-NFGAIL (IAP-N)], substituting asparagine (N) with phenylalanine (Syn-F/IAP-F), alanine (Syn-A/IAP-A), and glutamine (Syn-Q/IAP-Q). This allowed us to explore the effects of aromatic (π-system), aliphatic (hydrophobic), and hydrogen bonding effects with varying chain lengths on hydrogel formation. Our results reveal that Syn-N and Syn-Q exhibit MGC of 0.03 and 0.05 wt %, respectively, classifying them as super hydrogelators (MGC < 0.1 wt %). These values are 4.0-6.6-fold lower than Syn-F and Syn-A, with Syn-N demonstrating greater efficiency than Syn-Q. Similarly, IAP-N exhibited a substantial decrease in MGC by 8.75, 3.75, and 2.5 folds compared to IAP-A, IAP-F, and IAP-Q, respectively. Experimental evidence and molecular dynamic simulation suggest that -CO-NH2 of asparagine side chains effectively engaged in hydrogen bonding, thereby immobilizing water molecules at low gelator concentrations. Although glutamine shares similar -CO-NH2 functionality, its hydrogelation efficiency is less pronounced compared to asparagine, likely due to its longer alkyl chain, which may hinder the formation of a hydrogen bonding network in the self-assembled structure compared to asparagine-containing peptides. These findings offer valuable insights for designing efficient peptide hydrogelators or lowering MGCs by substituting amino acids with asparagine/glutamine in peptide sequences. Additionally, modifying peptide properties through asparagine/glutamine substitution could optimize hydrogel properties for specific applications.

Simultaneous electrochemical removal of three microcystin congeners and sulfamethoxazole in natural water

Microcystins (MCs), frequently detected in freshwater ecosystems, have raised significant human health and ecological concerns. New approaches are being developed to control and remove MCs. In this study, we examined factors influencing the efficacy of electrochemical oxidation as a means of control. Anode material (Pt/Ti, Ta2O5–IrO2/Ti, SnO2–SbO2/Ti, boron-doped diamond (BDD/Si), anode surface area ratios and solution volumes, initial pollutant concentrations, and the co-existing antibiotic sulfamethoxazole (SMX) were investigated. MCs and SMX were dissolved in filtered Taihu Lake water to simulate the natural aquatic environment. The results showed that non-active anodes, lower initial concentration of MC, larger surface area ratio of cathode to anode, and smaller ratio of reaction solution volume to anode surface area could promote the degradation target pollutants. Under optimal conditions in this study, the degradation rates of MC-LR, MC-YR, MC-RR, and SMX each reached more than 90% within 6 h, and the removal efficiency of MC-YR was the highest among three congeners. The effect of SMX on the degradation of MC congeners depended mainly on their concentration differences, such that when the initial concentration of SMX was one to two orders of magnitude lower than microcystin, the presence of SMX would promote the degradation of MCs. In contrast, when the initial concentration of SMX was higher than that of microcystin by approximately an order of magnitude, sulfamethoxazole would inhibit the degradation of MCs by between 4.6% and 24.5%. Ultra-high-performance liquid chromatography tandem mass spectrometry analysis revealed that the three MC congeners were electrochemically degraded through aromatic ring oxidation, alkene oxidation, and bond cleavage on the ADDA (3-amino-9-methoxy-2,6,8-trimethyl-10-phenyldeca-4,6-dienoic acid) side chain. Notably, the removal of MCs was accompanied by a decline in the hardness of the reaction water. This study provided insights into electrochemical degradation of microcystins and antibiotics in natural water, offering suggestions for its practical application.

Abiotic Acyl Transfer Cascades Driven by Aminoacyl Phosphate Esters and Self-Assembly.

Biochemical acyl transfer cascades, such as those initiated by the adenylation of carboxylic acids, are central to various biological processes, including protein synthesis and fatty acid metabolism. Designing cascade reactions in aqueous media remains challenging due to the need to control multiple, sequential reactions in a single pot and manage the stability of reactive intermediates. Herein, we developed abiotic cascades using aminoacyl phosphate esters, the synthetic counterparts of biological aminoacyl adenylates, to drive sequential chemical reactions and self-assembly in a single pot. We demonstrated that the structural elements of amino acid side chains (aromatic versus aliphatic) significantly influence the reactivity and half-lives of aminoacyl phosphate esters, ranging from hours to days. This behavior, in turn, affects the number of couplings we can achieve in the network and the self-assembly propensity of activated intermediate structures. The cascades are constructed using bifunctional peptide substrates featuring side chain nucleophiles. Specifically, aromatic amino acids facilitate the formation of transient thioesters, which preorganized into spherical aggregates and further couple into chimeric assemblies composed of esters and thioesters. In contrast, aliphatic amino acids, which lack the ability to form such structures, predominantly undergo hydrolysis, bypassing further transformations after thioester formation. Additionally, in mixtures containing multiple aminoacyl phosphate esters and peptide substrates, we achieved selective product formation by following a distinct pathway that favors subsequent reactions through reactivity changes and self-assembly. By coupling chemical reactions with molecules of varying reactivity time scales, we can drive multiple reaction clocks with distinct lifetimes and self-assembly dynamics, facilitating precise temporal and structural regulation.

An Integrated Module Performs Selective 'Online' Epoxidation in the Biosynthesis of the Antibiotic Mupirocin.

The delineation of the complex biosynthesis of the potent antibiotic mupirocin, which consists of a mixture of pseudomonic acids (PAs) isolated from Pseudomonas fluorescens NCIMB 10586, presents significant challenges, and the timing and mechanisms of several key transformations remain elusive. Particularly intriguing are the steps that process the linear backbone from the initial polyketide assembly phase to generate the first cyclic intermediate PA-B. These include epoxidation as well as incorporation of the tetrahydropyran (THP) ring and fatty acid side chain required for biological activity. Herein, we show that the mini-module MmpE performs a rare online (ACP-substrate) epoxidation and is integrated ('in-cis') into the polyketide synthase via a docking domain. A linear polyketide fragment with six asymmetric centres was synthesised using a convergent approach and used to demonstrate substrate flux via an atypical KS0 and a previously unannotated ACP (MmpE_ACP). MmpE_ACP-bound synthetic substrates were critical in demonstrating successful epoxidation in vitro by the purified MmpE oxidoreductase domain. Alongside feeding studies, these results confirm the timing as well as chain length dependence of this selective epoxidation. These mechanistic studies pinpoint the location and nature of the polyketide substrate prior to the key formation of the THP ring and esterification that generate PA-B.

Poly-Dha Sequences as Pro-polypeptides: An Original Mechanistic Postulate Leads to the Discovery of a Long-Acting Vasodilator KU04212.

The construction of polypeptides was revolutionized by Merrifield's solid-phase synthesis more than half a century ago. Herein, we explore a completely different approach to making peptides. We test an original mechanistic postulate wherein a single peptide made entirely of dehydroalanine (Dha) residues can give rise to regio- and stereodefined peptides by iterative conjugate addition of one- or two-electron nucleophiles. Each nucleophile appends a unique amino acid side chain to the peptide backbone. We show that side chain addition is not random. Side chains are added in one of two ways, in an electrophilicity-gated fashion (most cases) or in a substrate-directed manner, depending on the first nucleophile used in the synthesis. One peptide made in this series, KU04212, a first-in-class polyazole peptide, was found to reduce vascular length density (-17%; p < 0.05) and increase vessel diameter (124%; p < 0.001) in healthy day 6 chick embryos at 24 h post-single dose. It also rescued 75% of the embryos administered a 32-fold lethal dose of ischemia-inducing CoCl2 after 12 h and 12.5% of the embryos after 24 h. In comparison to three mechanistically distinct vasodilators, e.g., isosorbide mononitrate, amlodipine besylate, and prazosin, only KU04212 showed long-acting effects in vivo, making it an enticing lead for the treatment of ischemic disorders.

Monitoring Binding of Protamine with DNA Using Protein Charge Transfer Spectra.

In this work, novel intrinsic electronic absorption (250-400 nm) with a molar extinction coefficient of 752 M-1cm-1 at 250 nm, arising from photoinduced electron transfer involving charged amino acid side chains and the polypeptide backbone, along with luminescence (300-500 nm) with quantum yield of 0.011 from subsequent charge recombination, was observed in salmon sperm Protamine (PRM). The absorption of PRM was attributed to the previously identified Peptide Backbone-to-Side chain Charge Transfer (PBS-CT) from the polypeptide backbone to the abundant cationic headgroups of Arginine in PRM, while the luminescence was believed to originate from charge recombination within the charge-separated excited states of PRM. Remarkably, since Arg is the only charged residue in the PRM sequence, the PRM Protein Charge Transfer Spectra (ProCharTS) is both totally and uniquely Arg specific. Interestingly, the peak of PRM luminescence emission spectrum was independent of the excitation wavelength, unlike other proteins such as human serum albumin, displaying unconventional luminescence. Aggregation-induced effects on PRM absorbance and luminescence were ruled out, as both PRM absorbance and luminescence increase maintained linearity with increasing concentration in the 25-150 μM range. Nucleoprotamine complex formation, resulting from the binding of PRM with calf-thymus genomic DNA (gDNA), was monitored through increased scattering by the nucleoprotamine complex, a decrease in gDNA/PRM absorbance, a decrease in gDNA/PRM ellipticity, and shifts of nucleoprotamine complex band upon agarose gel electrophoresis. Upon binding with gDNA (700 μM base pair concentration), PRM ProCharTS absorbance at 260 nm decreased by 72%. This decrease was attributed to the formation and subsequent precipitation of nucleoprotamine complex upon PRM-gDNA binding. The application of ProCharTS absorbance to indirectly monitor DNA-protein binding in a label-free approach was thus demonstrated.

Introduction of the Trifluoropropionate Moiety into Aromatic Rings via Rhodium-Catalyzed Coupling of Arylboronic Acids with Diazoesters.

A catalytic method for the introduction of pharmaceutically meaningful fluorinated propionic acid side chains into aromatic compounds is presented. In the presence of rhodium catalyst [RhCp*Cl2]2, various arylboronic acids are efficiently coupled with an easy-to-access diazoester reagent to give perfluorinated derivatives of phenylpropionic acids, including top-selling profen drugs. The key advantage of this approach is that the pharmacophore is introduced as a whole and is compatible with various functionalities and drug discovery.