Structural Basis for Highly Efficient Production of Catechol Derivatives at Acidic pH by Tyrosinase from <i>Burkholderia thailandensis</i>

Son, Hyeoncheol Francis; Lee, Sanghyuk; Lee, Seul Hoo; Kim, Hyun; Hong, H.; Lee, Uk‐Jae; Lee, Pyung‐Gang; Kim, Byung Gee; Kim, Kyung-Jin

doi:10.1021/acscatal.8b02635

Cited by 20 publications

(23 citation statements)

References 34 publications

(59 reference statements)

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“…In contrast, most of the bacterial PPOs do not contain a signal peptide and the majority of them lack a C‐terminal domain [18,21] . However, some bacterial PPOs such as Burkholderia thailandensis ( Bt TYR), contain a C‐terminal domain similar to that in plant PPOs [33] . Fungal PPOs, like bacterial PPOs, mostly lack a signal peptide, but the majority of them contain a C‐terminal domain [12,21,22] .…”

Section: Type‐iii Copper Proteinsmentioning

confidence: 99%

Similar but Still Different: Which Amino Acid Residues Are Responsible for Varying Activities in Type‐III Copper Enzymes?

Kampatsikas

Rompel

2020

ChemBioChem

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Type‐III copper enzymes like polyphenol oxidases (PPOs) are ubiquitous among organisms and play a significant role in the formation of pigments. PPOs comprise different enzyme groups, including tyrosinases (TYRs) and catechol oxidases (COs). TYRs catalyze the o‐hydroxylation of monophenols and the oxidation of o‐diphenols to the corresponding o‐quinones (EC 1.14.18.1). In contrast, COs only catalyze the oxidation of o‐diphenols to the corresponding o‐quinones (EC 1.10.3.1). To date (August 2020), 102 PDB entries encompassing 18 different proteins from 16 organisms and several mutants have been reported, identifying key residues for tyrosinase activity. The structural similarity between TYRs and COs, especially within and around the active center, complicates the elucidation of their modes of action on a structural basis. However, mutagenesis studies illuminate residues that influence the two activities and show that crystallography on its own cannot elucidate the enzymatic activity mode. Several amino acid residues around the dicopper active center have been proposed to play an essential role in the two different activities. Herein, we critically review the role of all residues identified so far that putatively affect the two activities of PPOs.

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Section: Type‐iii Copper Proteinsmentioning

confidence: 99%

Similar but Still Different: Which Amino Acid Residues Are Responsible for Varying Activities in Type‐III Copper Enzymes?

Kampatsikas

Rompel

2020

ChemBioChem

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“…Recently, a tyrosinase from Burkholderia thaliandensis ( Bt Ty), which showed potent activity at low pH (ca. pH 4–5) with tetrameric structure, was found [ 40 ]. Since Bt Ty successfully converted daidzin (isoflavone glycoside), resveratrol (stilbene), and phloretin (chalcone) into their corresponding hydroxylated products, respectively, it was considered as a potential catalyst for the hydroxylation of kaempferol to produce quercetin as well as myricetin.…”

Section: Resultsmentioning

confidence: 99%

“…Along with the Bm Ty which had already been proven to show good activities on various polyphenols [ 40 ], the above-mentioned five tyrosinases were assessed for flavonol oxidation screening. However, all the tyrosinases except Bm Ty showed marginal activities for kaempferol and quercetin according to MBTH assay (quinone-detecting method) at pH 4 to 6 ( Figure 3 ).…”

Section: Resultsmentioning

confidence: 99%

“…However, all the tyrosinases except Bm Ty showed marginal activities for kaempferol and quercetin according to MBTH assay (quinone-detecting method) at pH 4 to 6 ( Figure 3 ). In the case of Bm Ty, because its optimal pH was around pH 8 [ 40 ] the flavonol-hydroxylating activity was relatively higher at slightly acidic pH (i.e., 6) than at acidic pH (i.e., 4 or 5). Since all the rest of the tyrosinases showed marginal activity, Bm Ty was chosen to perform further examination.…”

Section: Resultsmentioning

confidence: 99%

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Polyphenol-Hydroxylating Tyrosinase Activity under Acidic pH Enables Efficient Synthesis of Plant Catechols and Gallols

et al. 2021

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Tyrosinase is generally known as a melanin-forming enzyme, facilitating monooxygenation of phenols, oxidation of catechols into quinones, and finally generating biological melanin. As a homologous form of tyrosinase in plants, plant polyphenol oxidases perform the same oxidation reactions specifically toward plant polyphenols. Recent studies reported synthetic strategies for large scale preparation of hydroxylated plant polyphenols, using bacterial tyrosinases rather than plant polyphenol oxidase or other monooxygenases, by leveraging its robust monophenolase activity and broad substrate specificity. Herein, we report a novel synthesis of functional plant polyphenols, especially quercetin and myricetin from kaempferol, using screened bacterial tyrosinases. The critical bottleneck of the biocatalysis was identified as instability of the catechol and gallol under neutral and basic conditions. To overcome such instability of the products, the tyrosinase reaction proceeded under acidic conditions. Under mild acidic conditions supplemented with reducing agents, a bacterial tyrosinase from Bacillus megaterium (BmTy) displayed efficient consecutive two-step monophenolase activities producing quercetin and myricetin from kaempferol. Furthermore, the broad substrate specificity of BmTy toward diverse polyphenols enabled us to achieve the first biosynthesis of tricetin and 3′-hydroxyeriodictyol from apigenin and naringenin, respectively. These results suggest that microbial tyrosinase is a useful biocatalyst to prepare plant polyphenolic catechols and gallols with high productivity, which were hardly achieved by using other monooxygenases such as cytochrome P450s.

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“…[16] A similar pH dependence of the catalytic properties of tyrosinase was also recently reported for a Burkholderia thailandensis-derived tyrosinase. [36] In addition, the fusion enzyme was overexpressed in E. decreased the modification efficiency. In many other previous immobilization studies, the use of solid supports led to an increase in the steric hindrance between the enzyme and substrate.…”

Section: Discussionmentioning

confidence: 99%

Oriented in situ immobilization of a functional tyrosinase on microcrystalline cellulose effectively incorporates DOPA residues in bioengineered mussel adhesive protein

Kim

Bae

Shin

et al. 2021

Biotechnology Journal

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Background: Catechol-containing polymers such as mussel adhesive proteins (MAPs) are attractive as biocompatible adhesive biomaterials, and the catecholic amino acid 3,4-dihydroxyphenyl-L-alanine (DOPA) is considered a key molecule in underwater mussel adhesion. Tyrosinases can specifically convert tyrosine to DOPA without any cofactors. However, their catalytic properties still need to be adjusted to minimize unwanted DOPA oxidation via their diphenolase activity and catechol instability at neutral and basic pH values in the reaction products. Methods and Results:In this work, we constructed a novel functional tyrosinase, mTyr-CNK_CBM, by fusion of mTyr-CNK with a cellulose-binding motif (CBM) for oriented in situ immobilization on microcrystalline cellulose via the C-terminal CBM without any additional purification steps. mTyr-CNK_CBM showed optimal catalytic activity at pH 4.5-6.5 and room temperature and had a high monophenolase/diphenolase activity ratio (V max mono/V max di = 2.08 at pH 6 and 25 • C). mTyr-CNK_CBM exhibited 2.17fold higher (as a unimmobilized free enzyme) and similarly high (upon immobilization) in vitro DOPA modification of a bioengineered MAP compared to a commercially available mushroom tyrosinase. Moreover, the immobilized mTyr-CNK_CBM showed longterm storability and improved reusability.Conclusions: These results clearly demonstrate a strong potential for practical use of immobilized mTyr-CNK_CBM as a monophenol monooxygenase in preparing biocompatible DOPA-tethered biomaterials and other catechol-containing polymers.

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Structural Basis for Highly Efficient Production of Catechol Derivatives at Acidic pH by Tyrosinase from Burkholderia thailandensis

Cited by 20 publications

References 34 publications

Similar but Still Different: Which Amino Acid Residues Are Responsible for Varying Activities in Type‐III Copper Enzymes?

Similar but Still Different: Which Amino Acid Residues Are Responsible for Varying Activities in Type‐III Copper Enzymes?

Polyphenol-Hydroxylating Tyrosinase Activity under Acidic pH Enables Efficient Synthesis of Plant Catechols and Gallols

Oriented in situ immobilization of a functional tyrosinase on microcrystalline cellulose effectively incorporates DOPA residues in bioengineered mussel adhesive protein

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