Enzymatic Conversion of Unnatural Amino Acids by Yeast D‐Amino Acid Oxidase

Unnatural amino acids, particularly synthetic α‐amino acids, are becoming crucial tools for modern drug discovery research. In particular, this application requires enantiomerically pure isomers. In this work we report on the resolution of racemic mixtures of the amino acids d,l ‐naphthylalanine and d,l ‐naphthylglycine by using a natural enzyme, D ‐amino acid oxidase from the yeast Rhodotorula gracilis. A significant improvement of the bioconversion is obtained using a single‐point mutant enzyme designed by a rational approach. With this D ‐amino acid oxidase variant the complete resolution of all the unnatural amino acids tested was obtained: in this case, the bioconversion requires a shorter time and a lower amount of biocatalyst compared to the wild‐type enzyme. The simultaneous production of the corresponding α‐keto acid, a possible precursor of the amino acid in the L ‐form, improves the significance of the procedure.     

Amino acid oxidase‐catalysed resolution and Pictet–Spengler reaction towards chiral and rigid unnatural amino acids

BACKGROUND: The biosynthesis of structurally complex isoquinoline alkaloids and other natural products occurs via aromatic amino acids such as tyrosine, and chiral and rigid amino acids. These structures are also key building blocks of many active pharmaceutical ingredients. The aim of this work was the exploration of a rapid and straightforward route to chiral 6‐hydroxy‐1,2,3,4‐tetrahydroisoquinoline‐3‐carboxylic acid. RESULTS: The preparation of (S)‐meta‐tyrosine from racemic meta‐tyro‐ sine with aminoacidoxidase has been developed with ee > 99% and 88% yield. The combination of this resolution with a subsequent Pictet–Spengler reaction enables straightforward and versatile access to chiral (S)‐6‐hydroxy‐1,2,3,4‐tetrahydroisoquinoline‐3‐carboxylic acid in 30% yield. CONCLUSIONS: This new short chemoenzymatic route to (S)‐6‐hydroxy‐1,2,3,4‐tetrahydroisoquinoline‐3‐carboxylic acid from commercially available DL‐m‐tyrosine is more convenient than other chemical procedures and establishes a new link between the pool of easily accessible racemic aromatic amino acids and the corresponding chiral rigidified amino acids, which are of interest as structural elements of many active pharmaceutical ingredients. These results facilitate synthetic access to a range of active pharmaceutical ingredients and metabolites in chiral form from the oxidation of amino acids. This advances the opportunities to study the molecular interactions with enzymes, receptors and effectors more precisely than with the racemic forms. Copyright © 2007 Society of Chemical Industry     

RNA‐Directed Amino Acid Coupling as a Model Reaction for Primitive Coded Translation

The stereochemical theory claims that primitive coded translation initially occurred in the RNA world by RNA‐directed amino acid coupling. In this study, we show that the HIV Tat aptamer RNA is capable of recognizing two consecutive arginine residues within the Tat peptide, thus demonstrating how RNA might be able to position two amino acids for sequence‐specific coupling. We also show that this RNA can act as a template to accelerate the coupling of a single arginine residue to the N‐terminal arginine residue of a peptide primer. The results might have implications for our understanding of the origin of translation.     

An Aminocaprolactam Racemase from Ochrobactrum anthropi with Promiscuous Amino Acid Ester Racemase Activity

The kinetic resolution of amino acid esters (AAEs) is a useful synthetic strategy for the preparation of single‐enantiomer amino acids. The development of an enzymatic dynamic kinetic resolution (DKR) process for AAEs, which would give a theoretical yield of 100 % of the enantiopure product, would require an amino acid ester racemase (AAER); however, no such enzyme has been described. We have identified low AAER activity of 15 U mg^?1 in a homologue of a PLP‐dependent α‐amino ?‐caprolactam racemase (ACLR) from Ochrobactrum anthropi. We have determined the structure of this enzyme, OaACLR, to a resolution of 1.87 Å and, by using structure‐guided saturation mutagenesis, in combination with a colorimetric screen for AAER activity, we have identified a mutant, L293C, in which the promiscuous AAER activity of this enzyme towards l ‐phenylalanine methyl ester is improved 3.7‐fold.     

sp3‐sp2 C‐C Bond Formation via Brønsted Acid Trifluoromethanesulfonic Acid‐Catalyzed Direct Coupling Reaction of Alcohols and Alkenes

A novel and efficient trifluoromethanesulfonic acid‐catalyzed sp³‐sp² C C bond formation reaction through the direct coupling of alcohols with alkenes has been realized under mild conditions. The present protocol provides an attractive approach to a diverse range of polysubstituted olefins in good to excellent yields with high stereo‐ and regioselectivities.     

Palladium‐Catalyzed Synthesis of (Z)‐3‐Arylthioacrylic Acids and Thiochromenones

The three‐component reaction of aryl halides, sodium sulfide pentahydrate (Na₂S⋅5 H₂O), and propiolic acid in the presence of 2.5% bis(triphenylphosphine)palladium chloride [Pd(PPh₃)₂Cl₂], 5% 1,4‐bis(diphenylphosphino)butane (dppb) and 2 equivalents of 1,8‐diazabicycloundec‐7‐ene (DBU) produces stereoselectively (Z)‐3‐arylthioacrylic acids in good yields. A study of the reaction pathway suggested that the C S bond formation between aryl halides and Na₂S⋅5 H₂O proceeded first, and the resulting intermediate reacted with propiolic acid to produce the desired product. In addition, when the resulting product was treated with acid, the respective thiochromenones were formed in good yields.     

Identification and Characterization of D‐Succinylase,and a Proposed Enzymatic Method for D‐Amino Acid Synthesis

Chiral amino acids are important intermediates for the pharmaceutical industry. We have developed a novel one‐pot enzymatic method for D ‐amino acid synthesis by the dynamic kinetic resolution of N‐succinyl‐dl ‐amino acids using D ‐succinylase (DSA) and N‐succinylamino acid racemase (NSAR, EC 4.2.1.113). The DSA from Cupriavidus sp. P4‐10‐C, which hydrolyzes N‐succinyl‐D ‐amino acids enantioselectively to their corresponding D ‐amino acids, was identified for the first time by screening soil microorganisms. Subsequently, the DSA gene was cloned and overexpressed in Escherichia coli. DSA was shown to comprise two subunits with molecular masses of 26 kDa and 60 kDa. Additionally, the NSAR gene from Geobacillus stearothermphilus NCA1503, which racemizes N‐succinylamino acids, was also cloned and overexpressed in E. coli. The highly purified DSA and NSAR prepared from each recombinant E. coli were characterized and used for D ‐amino acid synthesis. A one‐pot enzymatic method converted 100 mM N‐succinyl‐dl ‐phenylalanine to D ‐phenylalanine in 91.1% conversion with 86.7% ee. This novel enzymatic method may be useful for the industrial production of many D ‐amino acids.

     

The Copper‐Catalyzed Oxidative N‐Acylation of Sulfoximines

An oxidative cross‐coupling reaction between aldehydes and sulfoximines involving dual C H/N H functionalization has been developed. This reaction process is facilitated by a simple copper catalyst (1 mol% loading) and tert‐butyl hydroperoxide (TBHP) as the oxidant and proceeds under mild reaction conditions to afford a series of valuable N‐acylated sulfoximine derivatives in excellent yields.     

Sulfonic Acid‐Catalyzed Autoxidative Carbon‐Carbon Coupling Reaction under Elevated Partial Pressure of Oxygen

An aerobic organocatalytic oxidative C C bond formation reaction of benzylic C H bonds with various C‐nucleophiles is described. The coupling reaction proceeds by simply stirring the substrates under elevated partial pressure of oxygen in the presence of a sulfonic acid catalyst at room temperature. Elevation of the pressure enables the reaction of a broad scope of nucleophile substrates otherwise showing poor reactivity at ambient pressure. The benzylic C H bonds of xanthene, acridanes, isochromane and related heterocycles could be functionalized with nucleophiles including ketones, 1,3‐dicarbonyl compounds and aldehydes. Electron‐rich arenes could be utilized as nucleophiles at elevated temperatures. The reactions are believed to proceed via autoxidation of the benzylic C H bonds to the hydroperoxides and subsequent nucleophilic substitution catalyzed by sulfonic acids.     

Lower Irritation Potential of Laureth‐3 Carboxylate Amino Acid Salt

Sodium laureth sulfate and amino acid type surfactants have been categorized as low‐skin irritation chemicals based on results of previous skin irritation tests. However, detergent‐induced skin irritation still occurs. Detergents with a low‐skin irritation effect are required, since the number of cosmetics for sensitive skin has increased. Therefore, an in vitro method of testing the safety of cosmetics for sensitive skin is required. Skin irritation by anionic surfactants was investigated to determine the effect of laureth‐3 carboxylate (polyoxyethylene lauryl carboxymethyl ether) amino acid salt on skin irritation. In addition, sodium laureth sulfate and amino acid type surfactants, regarded as low‐irritation surfactants, were also tested for skin irritation. The skin irritation effect of laureth‐3 carboxylate lysine salt (Surfactant 1 ), sodium laureth sulfate (Surfactant 2 ), and sodium N‐lauroyl glutamate (Surfactant 3 ) were investigated using a reconstructed human cultured epidermal model, LabCyte EPI‐MODEL24 6D. Cell viabilities of cultured epidermal cells exposed to Surfactant 1 (5.0 % aq.), Surfactant 2 , and Surfactant 3 were 82.0, 45.0 and 19.1 %, respectively. There were significant differences in cell viability upon exposure to 5.0 % aqueous test solutions of the three test chemicals. The results of the current investigation indicate that Surfactant 1 has a low skin‐irritation effect.     

Ruthenium‐Catalyzed Oxidative Homo‐Coupling of 2‐Arylpyridines

A ruthenium‐catalyzed oxidative homo‐coupling reaction of 2‐arylpyridines via C H activation was developed. The reaction could tolerate various functional groups on both the aryl and the pyridyl rings to afford a series of dimerized products with iron(III) chloride (FeCl₃) as a stoichiometric oxidant. A tentative mechanism was proposed for this oxidative C H/C H homo‐coupling.     

光学纯氨基酸的研究进展

氨基酸能参与多种多样的生命活动，此外它还是合成多种药物的重要中间体。大多数氨基酸都有一个或一个以上的手性中心，手性不同其生物活性也不同。因为生命活动对不同的氨基酸会有立体选择性，所以光学纯氨基酸在制药与食品行业有着非常重要的应用。本文综述了近些年国内外光学纯氨基酸的研究进展。     

Copper‐Catalyzed Cascade Reactions of Substituted 4‐Iodopyrazolecarbaldehydes with 1,2‐Phenylenediamines and 2‐Aminophenols

A new approach for the synthesis of novel annulated‐pyrazoles is presented. This protocol includes an intermolecular condensation followed by a copper‐mediated intramolecular C N or C O coupling reaction. The method is applied to a range of substituted 4‐iodopyrazolecarbaldehydes which react with 1,2‐phenylenediamines or 2‐aminophenols to yield substituted 2,4‐ or 1,4‐dihydrobenzo[b]pyrazolo[4,3‐e][1,4]diazepines or substituted‐2H‐ or 1H‐benzo[b]pyrazolo[3,4‐f][1,4]oxazepines, respectively.     

Ring-Opening Amino Esterification of Cyclic Ketones to Access Distal Amino Acid Derivatives

This study developed a method of synthesizing distal amino acid derivatives by the ring-opening reaction of cyclic ketones, following the amino esterification functionalization of both terminals. To achieve this goal, we performed the ring-opening reaction of cyclic ketones with an aminating reagent and alcohol under metal- and photocatalysis-free conditions in a single step. The method directly afforded distal amino acid derivatives bearing C4−C6, C8, and C12 carbon main chains, such as γ, δ, and ϵ-amino esters. The obtained amino esters were simply transformed into amino acids by hydrolysis.     

Highly Efficient Suzuki Coupling Reaction of α‐Chloroalkylidene‐β‐lactones and β‐Lactams with Organoboronic Acids

The activation of C Cl bond of (Z)‐α‐chloroalkylidene‐β‐lactones and (E)‐α‐chloroalkylidene‐β‐lactams via the Suzuki cross‐coupling reaction is reported in this paper. Alkyl, heteroaromatic, substituted phenyl‐ and alkenylboronic acids can be coupled with a wide variety of α‐chloroalkylidene‐β‐lactones and β‐lactams in excellent yields within a short period of time. The cross‐coupling reaction of optically active substrates leads to the optically active compounds without racemization of the corresponding chiral center.     

New Insight into the Structure–Activity Relationships of the Selective Excitatory Amino Acid Transporter Subtype 1 (EAAT1) Inhibitors UCPH‐101 and UCPH‐102

In the present study, we made further investigations on the structure–activity requirements of the selective excitatory amino acid transporter 1 (EAAT1) inhibitor, 2‐amino‐4‐(4‐methoxyphenyl)‐7‐(naphthalen‐1‐yl)‐5‐oxo‐5,6,7,8‐tetrahydro‐4H‐chromene‐3‐carbonitrile (UCPH‐101), by exploring 15 different substituents (R¹) at the 7‐position in combination with eight different substituents (R²) at the 4‐position. Among the 63 new analogues synthesized, we identified a number of compounds that unexpectedly displayed inhibitory activities at EAAT1 in light of understanding the structure–activity relationship (SAR) of this inhibitor class extracted from previous studies. Moreover, the nature of the R¹ and R² substituents were observed to contribute to the functional properties of the various analogues in additive and non‐additive ways. Finally, separation of the four stereoisomers of analogue 14 g (2‐amino‐4‐([1,1′‐biphenyl]‐4‐yl)‐3‐cyano‐7‐isopropyl‐5‐oxo‐5,6,7,8‐tetrahydro‐4H‐chromene) was carried out, and in agreement with a study of a related scaffold, the R configuration at C4 was found to be mandatory for inhibitory activity, while both the C7 diastereomers were found to be active as EAAT1 inhibitors. A study of the stereochemical stability of the four pure stereoisomers 14 g ‐ A – D showed that epimerization takes places at C7 via a ring‐opening, C?C bond rotation, ring‐closing mechanism.     

Palladium‐Catalyzed Cyclization Reactions of Acetylene‐Containing Amino Acids

Enantiomerically pure acetylene‐containing α‐amino acids were used as versatile starting materials for the synthesis of a variety of heterocycles via Pd‐mediated cyclization reactions. Depending on the protecting group strategy, both the carboxylate and the amine function of the amino acids could participate in the cyclizations, thus giving rise to oxygen heterocycles (α‐aminolactones) and nitrogen heterocycles (cyclic α‐amino acid derivatives), respectively. Beside the straightforward cyclization, cyclization/cross‐coupling reactions were also successfully carried out to provide the corresponding substituted cyclic amino acid derivatives.     

Palladium‐Catalysed Intramolecular Direct Arylation of 2‐Bromobenzenesulfonic Acid Derivatives

A palladium‐catalysed intramolecular direct arylation of 2‐bromobenzenesulfonic acid derivatives was found to proceed using 1 mol% of palladium acetate as the catalyst. The influence of the substituents on the phenol moiety of 2‐bromobenzenesulfonic acid phenyl esters reveals that electron‐donating substituents favour the reaction while electron‐withdrawing ones are unfavourable. The reactivity of sulfonamides was also studied and, in all cases, a selective activation at sp² C H vs. sp³ C H was observed. A sulfonamide bearing both phenyl and benzyl substituents on nitrogen gave selectively the six‐membered ring product.     

A New,Efficient and Recyclable Lanthanum(III) Oxide‐ Catalyzed C?N Cross‐Coupling

A new and efficient protocol for the C N cross‐coupling of aryl halides with heteroaromatic amines in the presence of lanthanum(III) oxide (10 mol%) as a recyclable catalyst, N,N′‐dimethylethylenediamine (DMEDA) (20 mol%) and potassium hydroxide (KOH) as a base in dimethyl sulfoxide (DMSO) at 110 °C has been developed. This inexpensive catalytic system is highly effective towards the amination of aryl halides with various nitrogen nucleophiles and is significantly tolerant towards other functional groups in the substrates.     

Evolving the N‐Terminal Domain of Pyrrolysyl‐tRNA Synthetase for Improved Incorporation of Noncanonical Amino Acids

By evolving the N‐terminal domain of Methanosarcina mazei pyrrolysyl‐tRNA synthetase (PylRS) that directly interacts with tRNA^Pyl, a mutant clone displaying improved amber‐suppression efficiency for the genetic incorporation of N^?‐(tert‐butoxycarbonyl)‐l ‐lysine threefold more than the wild type was identified. The identified mutations were R19H/H29R/T122S. Direct transfer of these mutations to two other PylRS mutants that were previously evolved for the genetic incorporation of N^?‐acetyl‐l ‐lysine and N^?‐(4‐azidobenzoxycarbonyl)‐l ‐δ,?‐dehydrolysine also improved the incorporation efficiency of these two noncanonical amino acids. As the three identified mutations were found in the N‐terminal domain of PylRS that was separated from its catalytic domain for charging tRNA^Pyl with a noncanonical amino acid, they could potentially be introduced to all other PylRS mutants to improve the incorporation efficiency of their corresponding noncanonical amino acids. Therefore, it represents a general strategy to optimize the pyrrolysine incorporation system‐based noncanonical amino‐acid mutagenesis.