1492-24-6Relevant articles and documents
Deracemization of unnatural amino acid: Homoalanine using d-amino acid oxidase and ω-transaminase
Seo, Young-Man,Mathew, Sam,Bea, Han-Seop,Khang, Yong-Ho,Lee, Sang-Hyeup,Kim, Byung-Gee,Yun, Hyungdon
, p. 2482 - 2485 (2012)
A deracemization method was developed to generate optically pure l-homoalanine from racemic homoalanine using d-amino acid oxidase and ω-transaminase. A whole cell reaction using a biphasic system converted 500 mM racemic homoalanine to 485 mM l-homoalanine (>99% ee). The Royal Society of Chemistry 2012.
Nonproteinogenic α-amino acid preparation using equilibrium shifted transamination
Li, Tao,Kootstra, Anna B.,Fotheringham, Ian G.
, p. 533 - 538 (2002)
Microbial α-transaminases such as tyrosine aminotransferase (TAT) and branched chain aminotransferase (BCAT) of Escherichia coli, are useful as industrial biocatalysts to prepare nonproteinogenic L-amino acids from α-keto acids and an amino donor. However, they typically yield only 50% product when L-glutamic acid, the preferred amino donor, is used due to accumulated 2-ketoglutaric acid. Accordingly, methods have been sought to increase the reaction yield by the recycle or removal of the keto acid by-product. In this report, we have investigated the biocatalytic coupling of δ-transamination with α-transamination to recycle 2-ketoglutaric acid, and thereby increase the yield of aminotransferase reaction products. Ornithine δ-aminotransferase (OAT) catalyses the reversible transfer of the δ-amino group of L-ornithine to 2-ketoglutaric acid forming L-glutamic acid semialdehyde and L-glutamic acid. The cyclisation of L-glutamic acid semialdehyde to form Δ1-pyrroline-5-carboxylate under physiological conditions, favours the reaction in the direction of L-glutamic acid formation. The Bacillus subtilis rocD gene encoding OAT was cloned and produced at high levels in E. coli. Combined cell extracts of separate E. coli strains overproducing OAT and E. coli tyrosine aminotransferase enabled the synthesis of L-2-aminobutyrate from 2-ketobutyric acid to reach a yield of 92% compared to 50% achievable by TAT alone. Similarly, combined extracts of strains overproducing OAT and E. coli branched-chain amino acid aminotransferase synthesised L-tert-leucine from trimethylpyruvic acid with a 73% yield compared to 31% with BCAT alone. The use of OAT as a general biocatalytic tool to achieve high yields in aminotransferase reactions is discussed.
Structure-guided engineering of: Pseudomonas dacunhae l-aspartate β-decarboxylase for l-homophenylalanine synthesis
Zhang, Min,Hu, Pengfei,Zheng, Yu-Cong,Zeng, Bu-Bing,Chen, Qi,Zhang, Zhi-Jun,Xu, Jian-He
, p. 13876 - 13879 (2020)
Structure-guided engineering of Pseudomonas dacunhael-aspartate β-decarboxylase (AspBDC) resulted in a double mutant (R37A/T382G) with remarkable 15400-fold improvement in specific activity reaching 216 mU mg-1, towards the target substrate 3(R)-benzyl-l-aspartate. A novel strategy for enzymatic synthesis of l-homophenylalanine was developed by using the variant as a biocatalyst affording 75% product yield within 12 h. Our results underscore the potential of engineered AspBDC for the biocatalytic synthesis of pharmaceutically relevant and value added unnatural l-amino acids.
The specificity and kinetic mechanism of branched-chain amino acid aminotransferase from Escherichia coli studied with a new improved coupled assay procedure and the enzyme's potential for biocatalysis
Yu, Xuejing,Wang, Xingguo,Engel, Paul C.
, p. 391 - 400 (2014)
Branched-chain amino acid aminotransferase (BCAT) plays a key role in the biosynthesis of hydrophobic amino acids (such as leucine, isoleucine and valine), and its substrate spectrum has not been fully explored or exploited owing to the inescapable restrictions of previous assays, which were mainly based on following the formation/consumption of the specific branched-chain substrates rather than the common amino group donor/acceptor. In our study, detailed measurements were made using a novel coupled assay, employing (R)-hydroxyglutarate dehydrogenase from Acidaminococcus fermentans as an auxiliary enzyme, to provide accurate and reliable kinetic constants. We show that Escherichia coli BCAT can be used for asymmetric synthesis of a range of non-natural amino acids such as l-norleucine, l-norvaline and l-neopentylglycine and compare the kinetic results with the results of molecular modelling. A full two-substrate steady-state kinetic study for several substrates yields results consistent with a bi-bi ping-pong mechanism, and detailed analysis of the kinetic constants indicates that, for good 2-oxoacid substrates, release of 2-oxoglutarate is much slower than release of the product amino acid during the transamination reaction. The latter is in fact rate-limiting under conditions of substrate saturation.
Radical SAM Activation of the B12-Independent Glycerol Dehydratase Results in Formation of 5′-Deoxy-5′-(methylthio) adenosine and Not 5′-Deoxyadenosine
Demick, Jonathan M.,Lanzilotta, William N.
, p. 440 - 442 (2011)
Activation of glycyl radical enzymes (GREs) by S-adenosylmethonine (AdoMet or SAM)-dependent enzymes has long been shown to proceed via the reductive cleavage of SAM. The AdoMet-dependent (or radical SAM) enzymes catalyze this reaction by using a [4Fe-4S] cluster to reductively cleave AdoMet to form a transient 5′deoxyadenosyl radical and methionine. This radical is then transferred to the GRE, and methionine and 5′deoxyadenosine are also formed. In contrast to this paradigm, we demonstrate that generation of a glycyl radical on the B12-independent glycerol dehydratase by the glycerol dehydratase activating enzyme results in formation of 5′deoxy- 5′(methylthio) adenosine and not 5′deoxyadenosine. This demonstrates for the first time that radical SAM activases are also capable of an alternative cleavage pathway for SAM.
Biocatalytic Cascade Reaction for the Asymmetric Synthesis of L- and D-Homoalanine
Silva, Marcus V. de M.,Costa, Ingrid C. R.,de Souza, Rodrigo O. M. A.,Bornscheuer, Uwe T.
, p. 407 - 411 (2019)
Unnatural amino acids attract growing attention for pharmaceutical applications as they are useful building blocks for the synthesis of a number of chiral drugs. Here, we describe a two-step enzymatic method for the asymmetric synthesis of homoalanine from L-methionine, a cheap and readily available natural amino acid. First, the enzyme L-methionine γ-lyase (METase), from Fusobacterium nucleatum, catalyzed the γ-elimination of L-methionine to 2-oxobutyrate. Second, an amino acid aminotransferase catalyzed the asymmetric conversion of 2-oxobutyrate to either L- or D-homoalanine. The L-branched chain amino acid aminotransferase from Escherichia coli (eBCAT), using L-glutamate as amino donor, produced L-homoalanine (32.5 % conv., 28 % y, 99 % ee) and the D-amino acid aminotransferase from Bacillus sp. (DATA) used D-alanine as amino donor to produce D-homoalanine (87.5 % conv., 69 % y, 90 % ee). Thus, this concept allows for the first time the synthesis of both enantiomers of this important unnatural amino acid.
Reductive Cleavage of Sulfoxide and Sulfone by Two Radical S-Adenosyl- l -methionine Enzymes
Mandalapu, Dhanaraju,Ji, Xinjian,Zhang, Qi
, p. 36 - 39 (2019)
Sulfoxides and sulfones are commonly found in nature as a result of thioether oxidation, whereas only a very few enzymes have been found to metabolize these compounds. Utilizing the strong reduction potential of the [4Fe-4S] cluster of radical S-adenosyl-l-methionine (SAM) enzymes, we herein report the first enzyme-catalyzed reductive cleavage of sulfoxide and sulfone. We show two radical SAM enzymes, tryptophan lyase NosL and the class C radical SAM methyltransferase NosN, are able to act on a sulfoxide SAHO and a sulfone SAHO2, both of which are structurally similar to SAM. NosL cleaves all of the three bonds (i.e., S-C(5′), S-C(γ), and S-O) connecting the sulfur center of SAHO, with a preference for S-C(5′) bond cleavage. Similar S-C cleavage activity was also found for SHAO2, but no S-O cleavage was observed. In contrast to NosL, NosN almost exclusively cleaves the S-C(5′) bonds of SAHO and SAHO2 with much higher efficiencies. Our study provides valuable insights into the [4Fe-4S] cluster-mediated reduction reactions and highlights the remarkable catalytic promiscuity of radical SAM enzymes.
Engineering of a novel biochemical pathway for the biosynthesis of L-2- aminobutyric acid in Escherichia coli K12
Fotheringham, Ian G.,Grinter, Nigel,Pantaleone, David P.,Senkpeil, Richard F.,Taylor, Paul P.
, p. 2209 - 2213 (1999)
L-2-Aminobutyric acid was synthesised in a transamination reaction from L-threonine and L-aspartic acid as substrates in a whole cell biotransformation using recombinant Escherichia coli K12. The cells contained the cloned genes tyrB, ilvA and alsS which respectively encode tyrosine aminotransferase of E. coli, threonine deaminase of E. coli and α- acetolactate synthase of B. subtilis 168. The 2-aminobutyric acid was produced by the action of the aminotransferase on 2-ketobutyrate and L- aspartate. The 2-ketobutyrate is generated in situ from L-threonine by the action of the deaminase, and the pyruvate by-product is eliminated by the acetolactate synthase. The concerted action of the three enzymes offers significant yield and purity advantages over the process using the transaminase alone with an eight to tenfold increase in the ratio of product to the major impurity.
Highly Stable Zr(IV)-Based Metal-Organic Frameworks for Chiral Separation in Reversed-Phase Liquid Chromatography
Jiang, Hong,Yang, Kuiwei,Zhao, Xiangxiang,Zhang, Wenqiang,Liu, Yan,Jiang, Jianwen,Cui, Yong
supporting information, p. 390 - 398 (2021/01/13)
Separation of racemic mixtures is of great importance and interest in chemistry and pharmacology. Porous materials including metal-organic frameworks (MOFs) have been widely explored as chiral stationary phases (CSPs) in chiral resolution. However, it remains a challenge to develop new CSPs for reversed-phase high-performance liquid chromatography (RP-HPLC), which is the most popular chromatographic mode and accounts for over 90% of all separations. Here we demonstrated for the first time that highly stable Zr-based MOFs can be efficient CSPs for RP-HPLC. By elaborately designing and synthesizing three tetracarboxylate ligands of enantiopure 1,1′-biphenyl-20-crown-6, we prepared three chiral porous Zr(IV)-MOFs with the framework formula [Zr6O4(OH)8(H2O)4(L)2]. They share the same flu topological structure but channels of different sizes and display excellent tolerance to water, acid, and base. Chiral crown ether moieties are periodically aligned within the framework channels, allowing for stereoselective recognition of guest molecules via supramolecular interactions. Under acidic aqueous eluent conditions, the Zr-MOF-packed HPLC columns provide high resolution, selectivity, and durability for the separation of a variety of model racemates, including unprotected and protected amino acids and N-containing drugs, which are comparable to or even superior to several commercial chiral columns for HPLC separation. DFT calculations suggest that the Zr-MOF provides a confined microenvironment for chiral crown ethers that dictates the separation selectivity.
Robustness, Entrainment, and Hybridization in Dissipative Molecular Networks, and the Origin of Life
Cafferty, Brian J.,Wong, Albert S. Y.,Semenov, Sergey N.,Belding, Lee,Gmür, Samira,Huck, Wilhelm T. S.,Whitesides, George M.
supporting information, p. 8289 - 8295 (2019/06/04)
How simple chemical reactions self-assembled into complex, robust networks at the origin of life is unknown. This general problem-self-assembly of dissipative molecular networks-is also important in understanding the growth of complexity from simplicity in molecular and biomolecular systems. Here, we describe how heterogeneity in the composition of a small network of oscillatory organic reactions can sustain (rather than stop) these oscillations, when homogeneity in their composition does not. Specifically, multiple reactants in an amide-forming network sustain oscillation when the environment (here, the space velocity) changes, while homogeneous networks-those with fewer reactants-do not. Remarkably, a mixture of two reactants of different structure-neither of which produces oscillations individually-oscillates when combined. These results demonstrate that molecular heterogeneity present in mixtures of reactants can promote rather than suppress complex behaviors.
