Are glutamate and asparagine necessary for tyrosinase activity of type-3 copper proteins?

Decker, Heinz; Solem, Even; Tuczek, Felix

doi:10.1016/j.ica.2017.11.031

Cited by 24 publications

(16 citation statements)

References 51 publications

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“…The TYR (+)‐larreatricin hydroxylase ( Lt LH), specialized in the synthesis of precursors of 8‐8′‐linked lignans specific to the creosote bush Larrea tridentata , integrates in between group 1 and group 2 enzymes (Figure ). Interestingly, both PPOs from Vitis vinifera that have been characterized regarding mono‐/diphenolase specificity, clearly belong to group 1 PPOs. Whereas Vv PPOg was described earlier to exhibit both mono‐ and diphenolase activity, Vv PPOcs‐3 was described to exhibit only diphenolase activity.…”

Section: Methodsmentioning

confidence: 99%

Catechol Oxidase versus Tyrosinase Classification Revisited by Site‐Directed Mutagenesis Studies

et al. 2019

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Catechol oxidases (COs) and tyrosinases (TYRs) are both polyphenol oxidases (PPOs) that catalyze the oxidation of ortho-diphenols to the corresponding quinones.B yt he official classification, only TYRs can also catalyze the hydroxylation of monophenols to ortho-diphenols.R esearchers have been trying to find the molecular reason for the mono-/diphenolase specificity for decades.H owever,t he hypotheses for the lacko fm onophenolase activity of plant COs are only based on crystal structures so far.T ot est these hypotheses,w ep erformed site-directed mutagenesis studies and phylogenetic analyses with dandelion PPOs offering high phylogenetic diversity,the results of which refute the structurebased hypotheses.W hile plant PPOs of phylogenetic group 2 solely exhibit diphenolase activity,plant PPOs of phylogenetic group 1u nexpectedly also show monophenolase activity.This finding sheds new light upon the molecular basis for mono-/ diphenol substrate specificity and challenges the current practice of generally naming plant PPOs as COs.Catechol oxidases (COs;E C1 .10.3.1) and tyrosinases (TYRs;E C1 .14.18.1) are ubiquitously distributed enzymes that are,for example,responsible for the undesired browning of damaged fruits and vegetables.A lthough they are of significant interest for biotechnological applications,t he molecular basis for their different activities-either only on ortho-diphenols (COs) or additionally on monophenols (TYRs;Scheme 1)-is still unknown. Scheme 1. Reactionsc atalyzed by catechol oxidases and tyrosinases.[*] Dr.

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Section: Methodsmentioning

confidence: 99%

Catechol Oxidase versus Tyrosinase Classification Revisited by Site‐Directed Mutagenesis Studies

et al. 2019

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“…Asn240 stabilizes cooperatively with a conserved glutamic acid (Glu235) a conserved water molecule via hydrogen bonding. The water molecule activated by Glu235 acts as a primary proton acceptor by abstracting a proton from the hydroxyl group of an incoming phenolic substrate, which is subsequently transferred to the carboxylic group of the conserved glutamic acid 59 . However, the replacement of Asn240 at the 1 st activity controller position cannot transform a TYR into a CO since Asn240Lys was still active on the monophenolic substrates phenol, tyrosol, tyramine and tyrosine.…”

Section: Kinetic Characterization Of Recombinant Jrppo1-wt Recombinamentioning

confidence: 99%

Conversion of walnut tyrosinase into a catechol oxidase by site directed mutagenesis

Panis

Kampatsikas

Bijelic

et al. 2020

Sci Rep

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polyphenol oxidases (ppos) comprise tyrosinases (tYRs) and catechol oxidases (cos), which catalyse the initial reactions in the biosynthesis of melanin. tYRs hydroxylate monophenolic (monophenolase activity) and oxidize diphenolic (diphenolase activity) substrates, whereas cos react only with diphenols. In order to elucidate the biochemical basis for the different reactions in PPOs, cDNA from walnut leaves was synthesized, the target gene encoding the latent walnut tyrosinase (jrPPO1) was cloned, and the enzyme was heterologously expressed in Escherichia coli. Mutations targeting the two activity controller residues (Asn240 and Leu244) as well as the gatekeeper residue (Phe260) were designed to impair monophenolase activity of jrPPO1. For the first time, monophenolase activity of jrPPO1 towards L-tyrosine was blocked in two double mutants (Asn240Lys/Leu244Arg and Asn240Thr/ Leu244Arg) while its diphenolase activity was partially preserved, thereby converting jrPPO1 into a CO. Kinetic data show that recombinant jrPPO1 resembles the natural enzyme, and spectrophotometric investigations proved that the copper content remains unaffected by the mutations. The results presented herein provide experimental evidence that a precisely tuned interplay between the amino acids located around the active center controls the substrate specificity and therewith the mono-versus diphenolase activity in the type-iii copper enzyme jrPPO1.Tyrosinases (TYRs), catechol oxidases (COs) and aurone synthases (AUSs) represent the polyphenol oxidase (PPO) family, which is an umbrella term for copper metalloenzymes 1-3 containing one type-III copper center. TYRs catalyse the hydroxylation of monophenols to o-diphenols (EC 1.14.18.1, monophenolase activity) as well as the subsequent oxidation of o-diphenols to their corresponding o-quinones (EC 1.10.3.1, diphenolase activity) 1,4 , whereas COs catalyse only the latter reaction, unable to react with monophenolic substrates (Fig. 1). AUSs participate in the formation of aurones from chalcone precursors and are involved in plant secondary metabolism 5,6 . Quinones produced by PPOs usually undergo non-enzymatic reactions, polymerize 7 and finally form melanin products 8,9 . PPOs occur in a broad spectrum of organisms, including archaea 10 , bacteria 11 , fungi 2 , plants 8 and animals 12,13 . In plants, they are believed to be involved in defence mechanisms associated with the formation of browning substances, which is triggered by mechanical damage or wounding 14 , while in animals their reaction products are responsible for coloring of skin, hair and eyes 13 .TYR from Juglans regia (walnut, jrPPO1) is expressed in vivo as a latent 66.8 kDa pro-enzyme consisting of three domains 15 : an N-terminal chloroplast transit peptide (~12 kDa) 15 , the catalytically active domain (~39 kDa) and the C-terminal domain (~16 kDa) that shields the entrance to the catalytic pocket and keeps the enzyme in a latent state. In vivo enzymatic activity is triggered by the removal of the C-terminal domain 16 . Alternati...

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“…Nevertheless, recently, some plant catechol oxidases have been described showing tyrosine hydroxylase activity [ 5 , 6 ], confirming that tyrosinases and catechol oxidases are very similar enzymes with subtle differences at the active site. They belong to the type 3 copper oxidases, as hemocyanins and even laccases [ 7 , 8 , 9 , 10 ]. In any case, after dopaquinone or any other o -quinone is formed by the oxidative action of these enzymes, the pathway progresses by a series of spontaneous reactions to lead to the final melanin, a structurally-ill-defined polymer [ 3 ].…”

Section: Introductionmentioning

confidence: 99%

“…All tyrosinases, including bacterial tyrosinases and plant polyphenol oxidases, contain a pair of copper ions at the flexible active site [ 44 ]. These enzymes belong to the type-3 copper-protein class, and they are currently being studied intensively [ 5 , 6 , 9 , 45 , 46 ]. These sites comprise six histidine residues that coordinate the two copper ions CuA and CuB.…”

Section: Introductionmentioning

confidence: 99%

“…This should be a huge advance to understand the features of the mammalian tyrosinase family and its respective role in the mammalian melanosome. It can also improve some remaining unconcluded questions in the mechanisms of action of the tyrosinase family, such as the differences among monophenol hydroxylase and o-diphenol oxidase activities or between tyrosinases and catechol oxidases looking for key residues determining the different affinities of both groups [ 5 , 6 , 8 , 9 , 45 ]. However, one of the most striking points from the structural point of view is the achievement of crystallized species of mammalian Trp1 containing Zn(II) instead of the expected Cu(II) [ 67 , 68 ].…”

Section: Introductionmentioning

confidence: 99%

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On the Metal Cofactor in the Tyrosinase Family

Solano

2018

IJMS

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The production of pigment in mammalian melanocytes requires the contribution of at least three melanogenic enzymes, tyrosinase and two other accessory enzymes called the tyrosinase-related proteins (Trp1 and Trp2), which regulate the type and amount of melanin. The last two proteins are paralogues to tyrosinase, and they appeared late in evolution by triplication of the tyrosinase gene. Tyrosinase is a copper-enzyme, and Trp2 is a zinc-enzyme. Trp1 has been more elusive, and the direct identification of its metal cofactor has never been achieved. However, due to its enzymatic activity and similarities with tyrosinase, it has been assumed as a copper-enzyme. Recently, recombinant human tyrosinase and Trp1 have been expressed in enough amounts to achieve for the first time their crystallization. Unexpectedly, it has been found that Trp1 contains a couple of Zn(II) at the active site. This review discusses data about the metal cofactor of tyrosinase and Trps. It points out differences in the studied models, and it proposes some possible points accounting for the apparent discrepancies currently appearing. Moreover, some proposals about the possible flexibility of the tyrosinase family to uptake copper or zinc are discussed.

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Are glutamate and asparagine necessary for tyrosinase activity of type-3 copper proteins?

Cited by 24 publications

References 51 publications

Catechol Oxidase versus Tyrosinase Classification Revisited by Site‐Directed Mutagenesis Studies

Catechol Oxidase versus Tyrosinase Classification Revisited by Site‐Directed Mutagenesis Studies

Conversion of walnut tyrosinase into a catechol oxidase by site directed mutagenesis

On the Metal Cofactor in the Tyrosinase Family

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