Production and characterization of a secreted, C‐terminally processed tyrosinase from the filamentous fungus Trichoderma reesei

Selinheimo, Emilia; Saloheimo, Markku; Ahola, Elina; Westerholm-Parvinen, Ann; Kalkkinen, Nisse; Büchert, Johanna; Kruus, Kristiina

doi:10.1111/j.1742-4658.2006.05429.x

Cited by 105 publications

(97 citation statements)

References 56 publications

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“…In addition, L -isomers of DOPA and tyrosine were much better substrates for B. megaterium tyrosinase than the corresponding D -isomers ( table 2 ). Very low oxidation of ocoumaric acid is in accordance with the literature regarding orthophenolic compounds, which are poor substrates for tyrosinases presumably because of steric hindrance [Selinheimo et al, 2006. Low oxidation activity towards catechol is remarkably different from most tyrosinases from other sources such as S. glaucesens and T. roseum [Kong et al, 2000;Lerch and Ettinger, 1972], but is comparable with reports for B. thuringiensis tyrosinase [Liu et al, 2004].…”

Section: Substrate Specificity and Inhibitorssupporting

confidence: 77%

“…This protein is also needed for the incorporation of copper and activation of the apotyrosinase. Recently, it was discovered that the filamentous fungus Trichoderma reesei has a secreted tyrosinase that is processed by cleavage of a 20-kDa peptide from its C-terminus [Selinheimo et al, 2006]. As the B. megaterium tyrosinase was successfully cloned in an active form in E. coli , it is assumed that there is no helper protein for this enzyme.…”

Section: Isolation and Identification Of A Tyrosinaseexpressing Bactementioning

confidence: 99%

“…[Liu et al, 2004], S. castaneoglobisporus (8.1 m M ) [Kohashi et al, 2004] or T. reesei (3 m M ) [Selinheimo et al, 2006] [Olivares et al, 2002].…”

Section: Mediummentioning

confidence: 99%

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Isolation, Cloning and Characterization of a Tyrosinase with Improved Activity in Organic Solvents from Bacillus megaterium

Shuster

Fishman²

2009

Microb Physiol

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A tyrosinase-expressing bacterium was isolated from soil, and extracellular enzymatic activity was induced by the presence of tyrosine and CuSO₄. Amplification of the 16S rDNA genes revealed a high similarity with Bacillus megaterium. The enzyme was over-expressed in Escherichia coli BL21 and purified using an affinity column. The tyrosinase was composed of 297 amino acids and was determined to be a monomer with a relative molecular mass of 31 kDa according to gel filtration. The K_m values for 3,4-dihydroxy-L-phenylalanine (L-DOPA) and L-tyrosine were 0.35 and 0.075 mM, respectively, and the k_cat/K_m values were 28.9·10³ and 32.9·10³ (s^–1·M^–1). The maximum activity for both monophenolase and diphenolase was observed at 50°C and pH 7.0. Enzymatic activity was enhanced in the presence of 10–50% water-miscible organic solvents, which included ethanol, methanol, 2-propanol and dimethyl sulfoxide (DMSO). The activity in 30% DMSO was 170% of the activity in water and the enantioselectivity towards L-DOPA decreased by 40%. The residual activity following an incubation period of 17 h in 0–70% methanol was constant. This newly isolated and characterized tyrosinase may have potential applications in organic synthesis due to its high activity and stability at typically denaturing conditions.

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Section: Substrate Specificity and Inhibitorssupporting

confidence: 77%

Section: Isolation and Identification Of A Tyrosinaseexpressing Bactementioning

confidence: 99%

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Isolation, Cloning and Characterization of a Tyrosinase with Improved Activity in Organic Solvents from Bacillus megaterium

Shuster

Fishman²

2009

Microb Physiol

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“…44 Saloheimo et al reported that the secreted filamentous fungus (Trichoderma reesei) tyrosinase ends with a Gly292 residue. 45,46 Besides these, Rompel et al recently reported that the mature mushroom tyrosinase (PPO4) was purified and its C-terminal end was determined to be Ser383 by MS analysis. 47 Interestingly, these activated tyrosinases have no C-terminal domain, and their C-terminal ends are located downstream of 4 amino acids after the YG motif, which is a domain border (linker region), although the amino acid sequences of this region are not conserved except the YG motif.…”

Section: Activation Of Pro-tyrosinase By Proteolytic Treatmentmentioning

confidence: 99%

Controlling Dicopper Protein Functions

Fujieda¹,

Itoh²

2016

Bulletin of the Chemical Society of Japan

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Maturation processes of dinuclear copper proteins such as tyrosinase, catechol oxidase, and hemocyanin have been a long-standing mystery in copper protein chemistry. Until now, several crystal structures have revealed that these copper proteins share a similar dinuclear copper active site, where each copper ion is ligated by three histidine imidazoles, and binds molecular oxygen in a side-on fashion to form a (¯-η 2 :η 2 -peroxido)dicopper(II) species not only as the dioxygen-adduct in oxy-hemocyanins but also as the key reactive intermediate for the hydroxylation of phenols to catechols (phenolase reaction) and the oxidation of catechols to o-quinones (catecholase reaction) in tyrosinases and catechol oxidases. Recently, we have succeeded in determining the high-resolution crystal structures of the recombinant pro-form of yellow koji mold tyrosinase to find the existence of a distinct C-terminal domain containing a CXXC unit, that is the common sequence motif of the copper chaperons. Thus, the C-terminal domain apparently acts as a copper chaperon, helping construction of the dinuclear copper active site of tyrosinase. Furthermore, we have found that the proteolytic cleavage of the C-terminal domain from the pro-form (inactive-form) of tyrosinase greatly enhances the tyrosinase activity, thus suggesting that the Cterminal domain also acts as a shielding domain to regulate the enzymatic activity. In fact, overall structure of the pro-form resembles the structure of one of the functional units of octopus hemocyanin (oxygen carrier protein), which also has a similar C-terminal domain prohibiting the monooxygenase activity. On the basis of these results together with the detailed kinetic and spectroscopic analyses, the maturation process of the dinuclear copper proteins is discussed to provide new insights into the regulation mechanism of the dicopper protein functions; dioxygen binding and activation. We have also succeeded in evolving phenolase activity from molluscan and arthropod hemocyanins by treating them with a hydrolytic enzyme or an acid, and demonstrated that the reaction mechanism of their phenolase activity is the same to that of tyrosinase itself, that is the electrophilic aromatic substitution mechanism. Furthermore, we have developed an artificial dicopper protein exhibiting catecholase activity using metallo-β-lactamase, a dinuclear zinc enzyme, as a metal binding platform.

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“…However, the fact that T. reesei has a laccase gene does not necessarily mean that it could degrade or modify lignin; the enzyme might have a different function. In a similar way to the laccase, a putative tyrosinase gene found in the genome sequence was cloned and expressed under the cbh1 promoter (Selinheimo et al, 2006). This tyrosinase has been found to have applications as a cross-linking agent in food processing (Selinheimo et al, 2007;Mattinen et al, 2008).…”

mentioning

confidence: 99%

The cargo and the transport system: secreted proteins and protein secretion in Trichoderma reesei (Hypocrea jecorina)

2012

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Trichoderma reesei (Hypocrea jecorina) is an efficient cell factory for protein production that is exploited by the enzyme industry. Yields of over 100 g secreted protein l "1 from industrial fermentations have been reported. In this review we discuss the spectrum of proteins secreted by T. reesei and the studies carried out on its protein secretion system. The major enzymes secreted by T. reesei under production conditions are those degrading plant polysaccharides, the most dominant ones being the major cellulases, as demonstrated by the 2D gel analysis of the secretome. According to genome analysis, T. reesei has fewer genes encoding enzymes involved in plant biomass degradation compared with other fungi with sequenced genomes. We also discuss other T. reesei secreted enzymes and proteins that have been studied, such as proteases, laccase, tyrosinase and hydrophobins. Investigation of the T. reesei secretion pathway has included molecular characterization of the pathway components functioning at different stages of the secretion process as well as analysis of the stress responses caused by impaired folding or trafficking in the pathway or by expression of heterologous proteins. Studies on the transcriptional regulation of the secretory pathway have revealed similarities, but also interesting differences, with other organisms, such as a different induction mechanism of the unfolded protein response and the repression of genes encoding secreted proteins under secretion stress conditions. IntroductionThe filamentous fungus Trichoderma reesei (Hypocrea jecorina) is used widely in the enzyme industry as a production organism (for reviews see Penttilä et al., 2004;Kubicek et al., 2009). It is primarily used for production of its native cellulolytic and hemicellulolytic enzymes, but also for production of heterologous proteins. In addition, T. reesei has served as an important model organism of fungal lignocellulose degradation. For example, most of the cellulase and hemicellulase enzymes were identified and characterized for the first time at genetic level in T. reesei (e.g. Shoemaker et al., 1983; Penttilä et al., 1986;Teeri et al., 1987), the molecular structures of these enzymes have been solved (e.g. Rouvinen et al., 1990;Divne et al., 1998) and the gene regulation mechanisms controlling the expression of these enzymes have been elucidated (Kubicek et al., 2009;Aro et al., 2005) from T. reesei. All the T. reesei strains used for research or protein production are derived from a single natural isolate. Extensive mutagenesis and screening programs have created a whole pedigree of strains with improved enzyme production properties. The genome of T. reesei was sequenced by the Joint Genome Institute (Martinez et al., 2008); the sequence is about 34 Mbp and it comprises about 9100 predicted gene models. Both the genome size and the gene number are smaller than that observed for other filamentous fungi. This is mostly explained by the fact that T. reesei has very little redundancy in its genome.During the last two dec...

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Production and characterization of a secreted, C‐terminally processed tyrosinase from the filamentous fungus Trichoderma reesei

Cited by 105 publications

References 56 publications

Isolation, Cloning and Characterization of a Tyrosinase with Improved Activity in Organic Solvents from <i>Bacillus megaterium</i>

Isolation, Cloning and Characterization of a Tyrosinase with Improved Activity in Organic Solvents from <i>Bacillus megaterium</i>

Controlling Dicopper Protein Functions

The cargo and the transport system: secreted proteins and protein secretion in Trichoderma reesei (Hypocrea jecorina)

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