Computer Modeling Explains the Structural Reasons for the Difference in Reactivity of Amine Transaminases Regarding Prochiral Methylketones

Amine transaminases (ATAs) are pyridoxal-5′-phosphate (PLP)-dependent enzymes that catalyze the transfer of an amino group from an amino donor to an aldehyde and/or ketone. In the past decade, the enzymatic reductive amination of prochiral ketones catalyzed by ATAs has attracted the attention of researchers, and more traditional chemical routes were replaced by enzymatic ones in industrial manufacturing. In the present work, the influence of the presence of an α,β-unsaturated system in a methylketone model substrate was investigated, using a set of five wild-type ATAs, the (R)-selective from Aspergillus terreus (Atr-TA) and Mycobacterium vanbaalenii (Mva-TA), the (S)-selective from Chromobacterium violaceum (Cvi-TA), Ruegeria pomeroyi (Rpo-TA), V. fluvialis (Vfl-TA) and an engineered variant of V. fluvialis (ATA-256 from Codexis). The high conversion rate (80 to 99%) and optical purity (78 to 99% ee) of both (R)- and (S)-ATAs for the substrate 1-phenyl-3-butanone, using isopropylamine (IPA) as an amino donor, were observed. However, the double bond in the α,β-position of 4-phenylbut-3-en-2-one dramatically reduced wild-type ATA reactivity, leading to conversions of <10% (without affecting the enantioselectivity). In contrast, the commercially engineered V. fluvialis variant, ATA-256, still enabled an 87% conversion, yielding a corresponding amine with >99% ee. Computational docking simulations showed the differences in orientation and intermolecular interactions in the active sites, providing insights to rationalize the observed experimental results.

Exploiting the Chiral Ligands of Bis(imidazolinyl)- and Bis(oxazolinyl)thiophenes—Synthesis and Application in Cu-Catalyzed Friedel–Crafts Asymmetric Alkylation

Five new C2-symmetric chiral ligands of 2,5-bis(imidazolinyl)thiophene (L1–L3) and 2,5-bis(oxazolinyl)thiophene (L4 and L5) were synthesized from thiophene-2,5-dicarboxylic acid (1) with enantiopure amino alcohols (4a–c) in excellent optical purity and chemical yield. The utility of these new chiral ligands for Friedel–Crafts asymmetric alkylation was explored. Subsequently, the optimized tridentate ligand L5 and Cu(OTf)2 catalyst (15 mol%) in toluene for 48 h promoted Friedel–Crafts asymmetric alkylation in moderate to good yields (up to 76%) and with good enantioselectivity (up to 81% ee). The bis(oxazolinyl)thiophene ligands were more potent than bis(imidazolinyl)thiophene analogues for the asymmetric induction of the Friedel–Crafts asymmetric alkylation.

Improved Synthesis of a Novel Biodegradable Tunable Micellar Polymer Based on Partially Hydrogenated Poly(β-malic Acid-co-benzyl Malate)

Poly(benzyl malate) (PBM), together with its derivatives, have been studied as nanocarriers for biomedical applications due to their superior biocompatibility and biodegradability. The acquisition of PBM is primarily from chemical routes, which could offer polymer-controlled molecular weight and a unique controllable morphology. Nowadays, the frequently used synthesis from L-aspartic acid gives an overall yield of 4.5%. In this work, a novel synthesis route with malic acid as the initiator was successfully designed and optimized, increasing the reaction yield up to 31.2%. Furthermore, a crystalline form of PBM (PBM-2) that polymerized from high optical purity benzyl-β-malolactonate (MLABn) was discovered during the optimization process. X-ray diffraction (XRD) patterns revealed that the crystalline PBM-2 had obvious diffraction peaks, demonstrating that its internal atoms were arranged in a more orderly manner and were different from the amorphous PBM-1 prepared from the racemic MLABn. The differential scanning calorimetry (DSC) curves and thermogravimetric curves elucidated the diverse thermal behaviors between PBM-1 and PBM-2. The degradation curves and scanning electron microscopy (SEM) images further demonstrated the biodegradability of PBM, which have different crystal structures. The hardness of PBM-2 implied the potential application in bone regeneration, while it resulted in the reduction of solubility when compared with PBM-1, which made it difficult to be dissolved and hydrogenated. The solution was therefore heated up to 75 °C to achieve benzyl deprotection, and a series of partially hydrogenated PBM was sequent prepared. Their optimal hydrogenation rates were screened to determine the optimal conditions for the formation of micelles suitable for drug-carrier applications. In summary, the synthesis route from malic acid facilitated the production of PBM for a shorter time and with a higher yield. The biodegradability, biosafety, mechanical properties, and adjustable hydrogenation widen the application of PBM with tunable properties as drug carriers.

Organocatalytic Asymmetric Michael Addition in Aqueous Media by a Hydrogen-Bonding Catalyst and Application for Inhibitors of GABAB Receptor

Catalysts based on (R, R)-1,2-diphenylethylenediamine are, as chiral organic catalysts, applied to the asymmetric Michael addition to α, β-unsaturated nitroalkenes under neutral conditions. The role of an aqueous medium for organic catalytic activity can be reversed concerning hydrophilic-hydrophobic function depending on the reaction conditions. In this study, to provide an environmentally friendly system, the thiourea-based catalyst substituted with 3,5-(CF3)2-Ph was used in water solvents. The hydrophobic effect of the substituent provided fast reaction, high chemical yield, and mirror-image selectivity. This reaction allowed the preparation of GABAB agonists in an optically pure manner. Additionally, GABA (γ-aminobutyric acid) analogs such as baclofen and phenibut were synthesized as R-type S-type with high optical purity.

Synthesis, Enantiomeric Resolution and Biological Evaluation of HIV Capsid Inhibition Activity for Racemic, (S)- and (R)-PF74

PF74 is a capsid-targeting inhibitor of HIV replication that effectively perturbs the highly sensitive viral uncoating process. A lack of information regarding the optical purity (enantiomeric excess) of the single stereogenic centre of PF74 has resulted in ambiguity as to the potency of different samples of this compound. Herein is described the synthesis of enantiomerically enriched (S)- and (R)-PF74 and further enrichment of the samples (≥98%) using chiral HPLC resolution. The biological activities of each enantiomer were then evaluated, which determined (S)-PF74 (IC50 1.5 µM) to be significantly more active than (R)-PF74 (IC50 19 µM). Computational docking studies were then conducted to rationalise this large discrepancy in activity, which indicated different binding conformations for each enantiomer. The binding energy of the conformation adopted by the more active (S)-PF74 (ΔG = −73.8 kcal/mol) was calculated to be more favourable than the conformation adopted by the less active (R)-enantiomer (ΔG = −55.8 kcal/mol) in agreement with experimental observations.

An ADH toolbox for raspberry ketone production from natural resources via a biocatalytic cascade

Raspberry ketone is a widely used flavor compound in food and cosmetic industry. Several processes for its biocatalytic production have already been described, but either with the use of genetically modified organisms (GMOs) or incomplete conversion of the variety of precursors that are available in nature. Such natural precursors are rhododendrol glycosides with different proportions of (R)- and (S)-rhododendrol depending on the origin. After hydrolysis of these rhododendrol glycosides, the formed rhododendrol enantiomers have to be oxidized to obtain the final product raspberry ketone. To be able to achieve a high conversion with different starting material, we assembled an alcohol dehydrogenase toolbox that can be accessed depending on the optical purity of the intermediate rhododendrol. This is demonstrated by converting racemic rhododendrol using a combination of (R)- and (S)-selective alcohol dehydrogenases together with a universal cofactor recycling system. Furthermore, we conducted a biocatalytic cascade reaction starting from naturally derived rhododendrol glycosides by the use of a glucosidase and an alcohol dehydrogenase to produce raspberry ketone in high yield. Key points • LB-ADH, LK-ADH and LS-ADH oxidize (R)-rhododendrol • RR-ADH and ADH1E oxidize (S)-rhododendrol • Raspberry ketone production via glucosidase and alcohol dehydrogenases from a toolbox Graphical abstract

Influence of Nitrogen Sources on D-Lactic Acid Biosynthesis by Sporolactobacillus laevolacticus DSM 442 Strain

The purpose of this study was to explore the possibility of replacing an expensive yeast extract contained in the fermentation medium for D-lactic acid (D-LA, R-lactic acid) biosynthesis with an alternative nitrogen source. The screening studies were conducted under stationary conditions and showed that pea seed hydrolysate was the most beneficial substrate in the process of D-LA biosynthesis by the strain Sporolactobacillus laevolacticus DSM 442 among the used inorganic and organic nitrogen sources, waste materials, food and agricultural products. After 96 h, 75.5 g/L D-LA was obtained in batch cultures in a medium containing pea seed hydrolysate, with an average productivity of 0.79 g/L/h, yield of 75.5%, and optical purity of 99.4%. In batch cultures fed once, in a medium with an analogous composition, 122.6 g/L D-LA was obtained after 120 h, and the average yield, productivity and optical purity were 87.6%, 1.021 g/L/h, and 99.6%, respectively. Moreover, the amount of D-LA obtained in the fermentation medium enriched with the above-mentioned cheap agricultural product was similar to the amounts obtained in the medium containing yeast extract in both stationary and bioreactor cultures. Our research shows that hydrolyzed pea seeds, which belong to the legume family, may be a promising nitrogen source for the production of D-LA on an industrial scale.

Effective D-lactic acid production from corncobs by simultaneous saccharification and fermentation using metabolically engineered Lactobacillus plantarum

A metabolically engineered Lactobacillus plantarum mutant, which could produce D-lactic acid from both glucose and xylose, was applied for the production of optically pure D-lactic acid from corncobs by simultaneous saccharification and fermentation (SSF). Using a corncob hydrolysate obtained by a combination of dilute acid treatment using 1.5% H2SO4 followed by enzymatic saccharification, the L. plantarum mutant completely assimilated both glucose and xylose in the corncob hydrolysate within 20 hours, resulting in the successful production of D-lactic acid with high optical purity in a batch culture. To improve the performance of D-lactic acid production from corncobs, SSF experiments from 100 to 250 g/L of acid-hydrolyzed corncobs using 1.5% H2SO4 were performed, and 49.7 to 101 g/L of D-lactic acid with 96.8-98.6% of optical purity was successfully produced. The D-lactic acid yield from corncobs (YL/C) was approx. 0.61 when 100-150 g/L of acid-hydrolyzed corncobs was used; however, the YL/C decreased to 0.49 as the concentration of acid-hydrolyzed corncobs because of insufficient acid hydrolysis of the corncobs. Therefore, by increasing the H2SO4 concentration to 3.5%, D-lactic acid production from corncobs significantly increased to 134 g/L with YL/C of 0.63 and 2.88 g/(L･h) of productivity from 250 g/L of acid-hydrolyzed corncobs.

Towards More Sustainable Materials for Geo-Environmental Engineering: The Case of Geogrids

Plastic materials are widely used in geotechnical engineering, especially as geosynthetics. The use of plastic-based products involves serious environmental risks caused by their degradation. Innovative research has been focusing on biodegradable polymers of natural origin, especially on poly(lactic acid) (PLA), to reduce the use of plastics. This study aims to explore the potentiality of biopolymers for the production of geogrids, measuring the chemical and mechanical characteristics of raw materials and of prototype samples, similar to those available on the market. First, chemical composition and optical purity were determined by hydrogen nuclear magnetic resonance (1H-NMR) and polarimetry. Furthermore, samples of uniaxial and biaxial geogrids were custom-molded using a professional 3D printer. Mechanical properties were measured both on the filament and on the prototype geogrids. The maximum tensile resistance was 6.76 kN/m for the neat-PLA filament and 10.14 kN/m for uniaxial prototype geogrids produced with PLA-based polymer mixed with titanium dioxide. PLA-based materials showed higher tensile properties than polypropylene (PP), the most common petroleum derivative. Conversely, such biomaterials seem to be more brittle and with scarce elongation rate respect PP. Nonetheless, these results are encouraging and can support the use of PLA-based materials for innovative biodegradable geosynthetics production, especially if used in combination with live plants.