Tyrosinase catalyzes the conversion of phenolic compounds into their quinone derivatives, which are precursors for the formation of melanin, a ubiquitous pigment in living organisms. Because of its importance for browning reactions in the food industry, the tyrosinase from the mushroom Agaricus bisporus has been investigated in depth. In previous studies the tyrosinase enzyme complex was shown to be a H(2)L(2) tetramer, but no clues were obtained of the identities of the subunits, their mode of association, and the 3D structure of the complex. Here we unravel this tetramer at the molecular level. Its 2.3 A resolution crystal structure is the first structure of the full fungal tyrosinase complex. The complex comprises two H subunits of ∼392 residues and two L subunits of ∼150 residues. The H subunit originates from the ppo3 gene and has a fold similar to other tyrosinases, but it is ∼100 residues larger. The L subunit appeared to be the product of orf239342 and has a lectin-like fold. The H subunit contains a binuclear copper-binding site in the deoxy-state, in which three histidine residues coordinate each copper ion. The side chains of these histidines have their orientation fixed by hydrogen bonds or, in the case of His85, by a thioether bridge with the side chain of Cys83. The specific tyrosinase inhibitor tropolone forms a pre-Michaelis complex with the enzyme. It binds near the binuclear copper site without directly coordinating the copper ions. The function of the ORF239342 subunits is not known. Carbohydrate binding sites identified in other lectins are not conserved in ORF239342, and the subunits are over 25 A away from the active site, making a role in activity unlikely. The structures explain how calcium ions stabilize the tetrameric state of the enzyme.

/pdf/crystal-structure-of-agaricus-bisporus-mushroom-tyrosinase-5egq24ti7m.pdf

Crystal structure of Agaricus bisporus mushroom tyrosinase: identity of the tetramer subunits and interaction with tropolone.

Tyrosinase plays a pivotal role in the synthesis of melanin pigment synthesis on skin utilizing tyrosine as a substrate. Melanin is responsible for the protection against harmful ultraviolet irradiation, which can cause significant pathological conditions, such as skin cancers. However, it can also create esthetic problems when accumulated as hyperpigmented spots. Various skin-whitening ingredients which inhibit tyrosinase activity have been identified. Some of them, especially ones with natural product origins, possess phenolic moiety and have been employed in cosmetic products. Semi-synthetic and synthetic inhibitors have also been developed under inspiration of the natural inhibitors yet some of which have no phenolic groups. In this review, tyrosinase inhibitors with natural, semi-synthetic and synthetic origins are listed up with their structures, activities and characteristics. Further, a recent report on the adverse effect of a natural melanin synthesis inhibitor which was included in skin-whitening cosmetics is also briefly discussed.

https://www.tandfonline.com/doi/pdf/10.3109/14756366.2015.1004058

Natural, semisynthetic and synthetic tyrosinase inhibitors

Melanogenesis is a biosynthetic pathway for the formation of the pigment melanin in human skin. A key enzyme, tyrosinase, catalyzes the first and only rate-limiting steps in melanogenesis, and the down-regulation of enzyme activity is the most reported method for the inhibition of melanogenesis. Because of the cosmetically important issue of hyperpigmentation, there is a big demand for melanogenesis inhibitors. This encourages researchers to seek potent melanogenesis inhibitors for cosmetic uses. This article reviews melanogenesis inhibitors that have been recently discovered from natural sources. The reaction mechanisms of the inhibitors on tyrosinase activity are also discussed.

Natural Melanogenesis Inhibitors Acting Through the Down-Regulation of Tyrosinase Activity

Natural ingredients have been used for centuries for skin care purposes. Nowadays, they are becoming more prevalent in formulations, due to consumers’ concerns about synthetic ingredients/chemical substances. The main benefits reported for plant extracts, used in skin care, include antioxidant and antimicrobial activities and tyrosinase inhibition effect. In this review, some examples of plants from Portuguese flora, whose extracts have shown good properties for skin care are presented. However, despite the known properties of plant extracts, few studies reported the development of formulations with them. More work in this field can be accomplished to meet consumer demand.

/pdf/main-benefits-and-applicability-of-plant-extracts-in-skin-5197q08jxb.pdf

Main Benefits and Applicability of Plant Extracts in Skin Care Products

Phytochemistry and biological properties of glabridin.

Skin-lightening products are commercially available for cosmetic purposes to obtain lighter skin complexion. Clinically, they are also used for treatment of hyperpigmentary disorders such as melasma, cafe au lait spot and solar lentigo. All of these target naturally melanin production, and many of the commonly used agents are known as competitive inhibitors of tyrosinase, one of the key enzymes in melanogenesis. In this review, we present an overview of commonly used skin-whitening ingredients that are commercialized, but we also hypothesize on other mechanisms that could be important targets to control skin pigmentation such as for example regulation of the adrenergic and glutaminergic signalling and also control of tetrahydrobiopterins in the human skin.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/j.1468-2494.2010.00616.x

The melanogenesis and mechanisms of skin-lightening agents--existing and new approaches

Synopsis

Objective

The need for effective ‘anti-ageing’ treatments, in particular for the management of photodamaged skin, prompted us to develop a novel method to identify new active ingredients. The model utilized a gene profiling study with corresponding connectivity mapping (Cmap) to identify novel anti-ageing compounds using all-trans retinoic acid (RA) as the lead compound due to its beneficial effect on photodamaged skin and skin firmness.



Method

A vehicle-controlled clinical study including nine healthy Caucasian female volunteers aged 57 ± 7 (mean ± SEM) exhibiting photodamage on their lower outer forearms was conducted. The volunteers applied RA once daily on photodamaged skin for 7 days, and biopsies were subjected to Affymetrix gene arrays. Connectivity mapping (Cmap), examining hundreds of gene expression profiles, was run on the gene signature of RA-treated photodamaged skin to identify small bioactive compounds.



Results

Affymetrix gene array identified 19 genes significantly differentially expressed after application of RA. Corresponding Cmap analysis revealed six natural bioactive compounds including N-acetyl aspartic acid (A-A-A) showing similar activity to RA on the differentially expressed genes identified.



Conclusion

Based on RA mimicking gene array activity, potential use within skincare on molecular size, safety assessment and sourcing, we identified the natural amino acid, A-A-A as a potential candidate to treat ageing skin.


Resume
Objectif

La demande croissante de traitements anti-âge efficaces, en particulier pour la traitement du photovieillissement, nous a incite a developper une methode pour identifier de nouveaux ingredients actifs. Le modele utilize la technologie de puces a ADN couplee au ‘connectivity mapping’ (Cmap) et permet d'identifier des composes anti-âge en utilisant l'acide retinoique comme molecule de reference en raison de son effet benefique sur la peau photo-âgee et notamment caracterisee par une perte de fermete.



Methodes

Une etude clinique a ete realisee sur neuf femmes volontaires, saines, de race blanche âgees de 57 ans ± 7, et presentant des signes de photovieillissement sur les avant-bras. La creme a l'acide retinoique a ete appliquee une fois par jour pendant sept jours. Sur les biopsies prelevees, l'analyse de l'expression genique a ete realisee a l'aide d'une puce a ADN. Le couplage avec la ‘connectivity mapping’ (Cmap) a permis d'identifier des composes potentiellement actifs mimant en partie l'activite de l'acide retinoique.



Resultats

L'etude a permis d'identifier une expression significative de 19 genes apres l'application d'acide retinoique. L'analyse en ‘connectivity mapping’ a revele six composes bioactifs naturels, y compris l'acide acetyl aspartique (A-A-A) montrant une activite similaire a l'acide retinoique.



Conclusion

Base sur un mimetisme d'activite de l'acide retinoique et en selectionnant des composes potentiellement actifs pour des soins cutanes en considerant leur poids moleculaire, l'evaluation de la securite, nous avons selectionne l'acide amine naturel, A-A-A comme un candidat potentiel pour traiter des signes du vieillissement cutane.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/ics.12251

The use of gene arrays and corresponding connectivity mapping (Cmap) to identify novel anti‐ageing ingredients

One of the most studied classes of skin-targeted compounds is retinoids. All-trans retinoic acid (RA) also referred to as tretinoin, and its synthetic or natural derivatives (retinoids) affect epidermal growth and differentiation 1. Synthetic retinoids are today used widely in the treatment of psoriasis and other disorders of keratinization 2. Also, acne has been widely treated with various forms of natural and synthetic retinoids for more than 40 years 3.

Later, retinoids has been used in the treatment for hyperpigmentation disorders such as post-inflammatory hyperpigmentation and melasma 4.

Separate from the effect of retinoids in skin disease, Kligman and Willis were the first to introduce retinoids for use as antiphotoaging agents 5. After its application, the authors noticed improvement in skin depigmentation and rejuvenation 6. Today, retinoids are extensively used for this indication 7,8.

Further studies have shown that RA's clinical effects include improvement in wrinkles, surface roughness, mottled pigmentation and skin appearance as a whole when used on photo-damaged skin 9–11. However, topical use of RA has also shown to develop erythema, scaling and burning/pruritus 12. Therefore, there is a constant thrive within the cosmetic industry to develop retinoid-like anti-ageing ingredients with significantly reduced side effects.

At a cellular level, retinoids are known to modulate the proliferation and differentiation of epidermal keratinocytes 13,14,14,15 by binding to nuclear retinoic acid receptors (RARs) and 9-cis-retinoic acid receptors (RXRs), which results in upregulation or downregulation of the transcription of target genes 16.

RA is the biologically active form of vitamin A (retinol). Endogenous production of RA by epidermal keratinocytes involves the uptake of preformed vitamin A from the surroundings followed by a series of metabolic activation steps involving retinol dehydrogenases (RDH1, RDH4, RDH10, RDH12 and DHRS9) and retinal dehydrogenases (RALDH1, RALDH2 and RALDH3) 17–19. DHRS9 specifically is encoding an enzyme that mediates conversion of retinol into RA.

Whilst endogenously produced or exogenously applied, RA is transported through the cytoplasm by specific intracellular retinoic-binding proteins (CRABP2) 8.

Once synthesized, the cellular levels of RA are controlled by several cytochrome P450-dependent enzymes (CYPs) – CYP26 A1, B1 and C1 – which metabolize RA into 4-hydroxy-RA, 4-oxo-RA and 18-OH-RA 20,21. CYP26B1 is known to be higher expressed than both CYP26A1 and CYP26C1 in human keratinocytes 19,22.

Over the last quarter century, 532 genes have been validated to be regulated by RA 23. In some cases, this control is direct, driven by a liganded RAR:RXR heterodimer bound to a DNA response element; in others, it is indirect, reflecting the actions of intermediate transcription factors, non-classical associations of receptors with other proteins or even more distant mechanisms.

Cell cultures have been studied extensively to characterize and prophesy the effect of retinoids on epidermal differentiation and growth 13,15,24–27.

In addition, a selection of genes using RT-PCR has been studied in full-thickness skin treated with retinoids 28,29.

Today, recent gene array techniques allow the characterization of the mRNA expression status of a large number of genes in cells or tissues after retinoid treatments with more than 170 studies conducted.

Some studies using gene arrays have also been carried out on reconstructed epidermis and human epidermis in vivo 30.

However, to our knowledge, gene array analysis following the treatment of RA on organoculture of full-thickness skin has not been investigated earlier.

The aim of the present study was to determine whether it is possible to use an organoculture of full-thickness skin as a preclinical model to evaluate modulation of gene expression profile following topical application of a commercially available retinoid cream.

https://www.onlinelibrary.wiley.com/doi/pdf/10.1111/ics.12121

M. J. Olsson

Papers

The melanogenesis and mechanisms of skin-lightening agents--existing and new approaches

The use of gene arrays and corresponding connectivity mapping (Cmap) to identify novel anti‐ageing ingredients

Transcriptional changes in organoculture of full-thickness human skin following topical application of all-trans retinoic acid