1133-63-7Relevant academic research and scientific papers
Flavoenzyme-mediated Regioselective Aromatic Hydroxylation with Coenzyme Biomimetics
Guarneri, Alice,Westphal, Adrie H.,Leertouwer, Jos,Lunsonga, Joy,Franssen, Maurice C. R.,Opperman, Diederik J.,Hollmann, Frank,van Berkel, Willem J. H.,Paul, Caroline E.
, p. 1368 - 1375 (2020/02/04)
Regioselective aromatic hydroxylation is desirable for the production of valuable compounds. External flavin-containing monooxygenases activate and selectively incorporate an oxygen atom in phenolic compounds through flavin reduction by the nicotinamide adenine dinucleotide coenzyme, and subsequent reaction with molecular oxygen. This study provides the proof of principle of flavoenzyme-catalyzed selective aromatic hydroxylation with coenzyme biomimetics. The carbamoylmethyl-substituted biomimetic in particular affords full conversion in less than two hours for the selective hydroxylation of 5 mM 3- and 4-hydroxybenzoates, displaying similar rates as with NADH, achieving a 10 mM/h enzymatic conversion of the medicinal product gentisate. This biomimetic appears to generate less uncoupling of hydroxylation that typically leads to undesired hydrogen peroxide. Therefore, we show these flavoenzymes have the potential to be applied in combination with biomimetics.
Altering 2-Hydroxybiphenyl 3-Monooxygenase Regioselectivity by Protein Engineering for the Production of a New Antioxidant
Bregman-Cohen, Almog,Deri, Batel,Maimon, Shiran,Pazy, Yael,Fishman, Ayelet
, p. 583 - 590 (2018/02/13)
2-Hydroxybiphenyl 3-monooxygenase is a flavin-containing NADH-dependent aromatic hydroxylase that oxidizes a broad range of 2-substituted phenols. In order to modulate its activity and selectivity, several residues in the active site pocket were investigated by saturation mutagenesis. Variant M321A demonstrated altered regioselectivity by oxidizing 3-hydroxybiphenyl for the first time, thus enabling the production of a new antioxidant, 3,4-dihydroxybiphenyl, with similar ferric reducing capacity to the well-studied piceatannol. The crystal structure of M321A was determined (2.78 ?), and molecular docking of the 3-substituted phenol provided a rational explanation for the altered regioselectivity. Furthermore, HbpA was found to possess pro-S enantioselectivity towards the production of several chiral sulfoxides, whereas variant M321F exhibited improved enantioselectivity. Based on the biochemical characterization of several mutants, it was suggested that Trp97 stabilized the substrate in the active site, Met223 was involved in NADH entrance or binding to the active site, and Pro320 might facilitate FAD movement.
Conversion of Simple Cyclohexanones into Catechols
Liang, Yu-Feng,Li, Xinyao,Wang, Xiaoyang,Zou, Miancheng,Tang, Conghui,Liang, Yujie,Song, Song,Jiao, Ning
supporting information, p. 12271 - 12277 (2016/09/28)
A novel I2-catalyzed direct conversion of cyclohexanones to substituted catechols under mild and simple conditions has been described. This novel transformation is remarkable with the multiple oxygenation and dehydrogenative aromatization processes enabled just by using DMSO as the solvent, oxidant, and oxygen source. This metal-free and simple system demonstrates a versatile protocol for the synthesis of highly valuable substituted catechols and therefore streamlines the synthesis and modification of biologically important molecules for drug discovery.
Structures of the Apo and FAD-Bound Forms of 2-Hydroxybiphenyl 3-monooxygenase (HbpA) Locate Activity Hotspots Identified by Using Directed Evolution
Jensen, Chantel N.,Mielke, Tamara,Farrugia, Joseph E.,Frank, Annika,Man, Henry,Hart, Sam,Turkenburg, Johan P.,Grogan, Gideon
, p. 968 - 976 (2015/04/14)
The FAD-dependent monooxygenase HbpA from Pseudomonas azelaica HBP1 catalyses the hydroxylation of 2-hydroxybiphenyl (2HBP) to 2,3-dihydroxybiphenyl (23DHBP). HbpA has been used extensively as a model for studying flavoprotein hydroxylases under process conditions, and has also been subjected to directed-evolution experiments that altered its catalytic properties. The structure of HbpA has been determined in its apo and FAD-complex forms to resolutions of 2.76 and 2.03 ?, respectively. Comparisons of the HbpA structure with those of homologues, in conjunction with a model of the reaction product in the active site, reveal His48 as the most likely acid/base residue to be involved in the hydroxylation mechanism. Mutation of His48 to Ala resulted in an inactive enzyme. The structures of HbpA also provide evidence that mutants achieved by directed evolution that altered activity are comparatively remote from the substrate-binding site.
A crystal structure of 2-hydroxybiphenyl 3-monooxygenase with bound substrate provides insights into the enzymatic mechanism
Kanteev, Margarita,Bregman-Cohen, Almog,Deri, Batel,Shahar, Anat,Adir, Noam,Fishman, Ayelet
, p. 1906 - 1913 (2015/10/28)
2-Hydroxybiphenyl 3-monooxygenase (HbpA) is an FAD dependent monooxygenase which catalyzes the ortho-hydroxylation of a broad range of 2-substituted phenols in the presence of NADH and molecular oxygen. We have determined the structure of HbpA from the soil bacterium Pseudomonas azelaica HBP1 with bound 2-hydroxybiphenyl, as well as several variants, at a resolution of 2.3-2.5 ? to investigate structure function correlations of the enzyme. An observed hydrogen bond between 2-hydroxybiphenyl and His48 in the active site confirmed the previously suggested role of this residue in substrate deprotonation. The entrance to the active site was confirmed by generating variant G255F which exhibited only 7% of the wild-type's specific activity of product formation, suggesting inhibition of substrate entrance into the active site by the large aromatic residue. Residue Arg242 is suggested to facilitate FAD movement and reduction as was previously reported in studies on the homologous protein para-hydroxybenzoate hydroxylase. In addition, it is suggested that Trp225, which is located in the active site, facilitates proper substrate entrance into the binding pocket in contrast to aklavinone-11-hydroxylase and para-hydroxybenzoate hydroxylase in which a residue at a similar position is responsible for substrate deprotonation. Structure function correlations described in this work will aid in the design of variants with improved activity and altered selectivity for potential industrial applications.
Arene cis-Diol Dehydrogenase-Catalysed Regio- and Stereoselective Oxidation of Arene-, Cycloalkane- and Cycloalkene-cis-diols to Yield Catechols and Chiral α-Ketols
Boyd, Derek R.,Sharma, Narain D.,Berberian, M. Victoria,Cleij, Marcel,Hardacre, Christopher,Ljubez, Vera,McConville, Gareth,Stevenson, Paul J.,Kulakov, Leonid A.,Allen, Christopher C. R.
supporting information, p. 1881 - 1894 (2015/06/02)
Benzene cis-diol dehydrogenase and naphthalene cis-diol dehydrogenase enzymes, expressed in Pseudomonas putida wild-type and Escherichia coli recombinant strains, were used to investigate regioselectivity and stereoselectivity during dehydrogenations of arene, cyclic alkane and cyclic alkene vicinal cis-diols. The dehydrogenase-catalysed production of enantiopure cis-diols, α-ketols and catechols, using benzene cis-diol dehydrogenase and naphthalene cis-diol dehydrogenase, involved both kinetic resolution and asymmetric synthesis methods. The chemoenzymatic production and applications of catechol bioproducts in synthesis were investigated.
Regioselective biocatalytic aromatic hydroxylation in a gas-liquid multiphase tube-in-tube reactor
Tomaszewski, Bartlomiej,Lloyd, Richard C.,Warr, Antony J.,Buehler, Katja,Schmid, Andreas
, p. 2567 - 2576 (2015/04/14)
Microreactors provide higher mass transfer rates than do conventional batch reactors. A tube-in-tube microreactor was used for the NADH-dependent in vitro conversion of 2-hydroxybiphenyl to 3-phenylcatechol that was catalysed by 2-hydroxybiphenyl 3-monooxygenase. A biphasic reaction system allowed high substrate loadings, whereas the microreactor ensured excellent mass transfer rates between the organic and aqueous phases. Oxygen was supplied continuously by membrane aeration across the whole reaction compartment. The productivities achieved in the tube-in-tube microreactor were 38 times higher than those in previously described batch reactors and almost 4 times higher than for the same reaction in a microreactor in which aqueous, organic, and air phases were delivered through consecutive segments. This set-up is a promising concept for oxygen-dependent biocatalytic reactions in microreactors and is developing as a basis for applications in gram-scale organic biosyntheses. Flow power: A tube-in-tube reactor is presented for a gas-dependent biocatalytic reaction, overcoming typical limitations such as mass transfer, product and substrate inhibition, and challenges with gas delivery with productivities superior to standard batch reactors or conventional microreactors.
Biocatalytic production of catechols using a high pressure tube-in-tube segmented flow microreactor
Tomaszewski, Bartlomiej,Schmid, Andreas,Buehler, Katja
, p. 1516 - 1526 (2015/02/19)
This study reports the synthesis of 3-phenylcatechol at the preparative scale using a continuous segmented flow tube-in-tube reactor (TiTR). 2-Hydroxybiphenyl 3-monooxygenase (HbpA) was applied as a biocatalyst for the hydroxylation reaction, which is dependent on the substrate 2-hydroxybiphenyl, NADH, and oxygen. While the regeneration of the cofactor NADH was guaranteed by formate dehydrogenase (FDH), oxygen was supplied via the membrane surface from the outside of the reactor system. The oxygen transfer rate through the membrane of the TiTR was determined to be 24 μmol O2 min-1 mL-1 emphasizing the potential of the TiTR as promising technology for realizing gas-dependent enzymatic reactions. Residence time and total turnover number have been identified as key limiting parameters. It was possible to scale-up this system by extending the TiTR by additional residence time units. This allowed synthesis of 1 g of 3-phenylcatechol at a high space time yield of 14.5 g L-1 h-1.
Synthetic cascades are enabled by combining biocatalysts with artificial metalloenzymes
Koehler,Wilson,Duerrenberger,Ghislieri,Churakova,Quinto,Knoerr,Haeussinger,Hollmann,Turner,Ward
, p. 93 - 99 (2013/08/24)
Enzymatic catalysis and homogeneous catalysis offer complementary means to address synthetic challenges, both in chemistry and in biology. Despite its attractiveness, the implementation of concurrent cascade reactions that combine an organometallic catalyst with an enzyme has proven challenging because of the mutual inactivation of both catalysts. To address this, we show that incorporation of a d 6 -piano stool complex within a host protein affords an artificial transfer hydrogenase (ATHase) that is fully compatible with and complementary to natural enzymes, thus enabling efficient concurrent tandem catalysis. To illustrate the generality of the approach, the ATHase was combined with various NADH-, FAD- and haem-dependent enzymes, resulting in orthogonal redox cascades. Up to three enzymes were integrated in the cascade and combined with the ATHase with a view to achieving (i) a double stereoselective amine deracemization, (ii) a horseradish peroxidase-coupled readout of the transfer hydrogenase activity towards its genetic optimization, (iii) the formation of L-pipecolic acid from L-lysine and (iv) regeneration of NADH to promote a monooxygenase-catalysed oxyfunctionalization reaction.
A flavin-dependent monooxygenase from Mycobacterium tuberculosis involved in cholesterol catabolism
Dresen, Carola,Lin, Leo Y.-C.,D'Angelo, Igor,Tocheva, Elitza I.,Strynadka, Natalie,Eltis, Lindsay D.
experimental part, p. 22264 - 22275 (2011/04/17)
Mycobacterium tuberculosis (Mtb) and Rhodococcus jostii RHA1 have similar cholesterol catabolic pathways. This pathway contributes to the pathogenicity of Mtb. The hsaAB cholesterol catabolic genes have been predicted to encode the oxygenase and reductase, respectively, of a flavin-dependent mono-oxygenase that hydroxylates 3-hydroxy-9,10-seconandrost-1,3,5(10)-triene-9,17-dione (3-HSA) to a catechol. An hsaA deletion mutant of RHA1 did not grow on cholesterol but transformed the latter to 3-HSA and related metabolites in which each of the two keto groups was reduced: 3,9-dihydroxy-9,10-seconandrost-1,3,5(10)-triene-17- one (3,9-DHSA) and 3,17-dihydroxy-9,10-seconandrost-1,3,5(10)-triene-9-one (3,17-DHSA). Purified 3-hydroxy-9,10-seconandrost-1,3,5(10)-triene-9,17-dione 4-hydroxylase (HsaAB) from Mtb had higher specificity for 3-HSA than for 3,17-DHSA (apparent kcat/Km = 1000 ± 100 M -1 s-1 versus 700 ± 100 M-1 s -1). However, 3,9-DHSA was a poorer substrate than 3-hydroxybiphenyl (apparent kcat/Km = 80 ± 40 M-1 s -1). In the presence of 3-HSA the Kmapp for O2 was 100 ± 10 μM. The crystal structure of HsaA to 2.5-A resolution revealed that the enzyme has the same fold, flavin-binding site, and catalytic residues as p-hydroxyphenyl acetate hydroxylase. However, HsaA has a much larger phenol-binding site, consistent with the enzyme's substrate specificity. In addition, a second crystal form of HsaA revealed that a C-terminal flap (Val367-Val394) could adopt two conformations differing by a rigid body rotation of 25° around Arg 366. This rotation appears to gate the likely flavin entrance to the active site. In docking studies with 3-HSA and flavin, the closed conformation provided a rationale for the enzyme's substrate specificity. Overall, the structural and functional data establish the physiological role of HsaAB and provide a basis to further investigate an important class of monooxygenases as well as the bacterial catabolism of steroids.
