Raman Characterization of Fungal DHN and DOPA Melanin Biosynthesis Pathways

Fungal melanins represent a resource for important breakthroughs in industry and medicine, but the characterization of their composition, synthesis, and structure is not well understood. Raman spectroscopy is a powerful tool for the elucidation of molecular composition and structure. In this work, we characterize the Raman spectra of wild-type Aspergillus fumigatus and Cryptococcus neoformans and their melanin biosynthetic mutants and provide a rough “map” of the DHN (A. fumigatus) and DOPA (C. neoformans) melanin biosynthetic pathways. We compare this map to the Raman spectral data of Aspergillus nidulans wild-type and melanin biosynthetic mutants obtained from a previous study. We find that the fully polymerized A. nidulans melanin cannot be classified according to the DOPA pathway; nor can it be solely classified according to the DHN pathway, consistent with mutational analysis and chemical inhibition studies. Our approach points the way forward for an increased understanding of, and methodology for, investigating fungal melanins.

Characterization of The Physicochemical Properties, Antioxidant Activity, And Antiproliferative Activity of Natural Melanin From S. Reiliana

This study aimed to characterize the physicochemical properties and stability of L-25 melanin extracted from Sporisorium reilianum (S. reiliana). The results showed that the maximum absorption wavelength of melanin was 215 nm. Reducing agents, heat, light, microwaving, oxidants, and common food additives did not affect the melanin. Additionally, it has a good metal stability except Mn2+. The IR spectra revealed the presence of O-H, N-H, C=O, and C=C bonds as well as carboxyl, alcohol hydroxyl, and phenolic hydroxyl groups and a pyran ring. L-25 melanin could be defined as DL-hydroxy phenylalanine (DOPA)-melanin. The antioxidant and antiproliferative were also measured. The melanin has a specific stability and high antioxidant activity, including a strong DPPH free radical scavenging ability, and protected damaged HepG2 cells by reducing reactive oxygen species, malondialdehyde, and lactate dehydrogenase content. In conclusion, S. reilianum represents a novel source of melanin, that could be applied to health food or food additives. Our results show that melanin from S. reilianum is a natural pigment with good stability that has a great prospect of development and application, providing a theoretical basis and methods for its further processing and development as a functional food.

Microbial Production and Characterization of 3, 4-dihydroxy-L Phenylalanine Mediated an Extracellular Heterogenic Eumelanin Pigment From Marine Actinomycetes Prauserella Sp. MAPPL 017A

Melanin is one of the natural biological pigment and it is derived from the enzyme tyrosinase by the oxidation of an amino acid tyrosine. The enzyme tyrosinase [EC 1.14.18.1] is a type-3 copper protein family found large amount in microorganisms and human beings involved in the biosynthesis of melanin and other polyphenolic compounds. Melanin plays an important role in all living creatures such as photo protectants against UV radiation, antioxidant, metal adsorption, electrical conductivity, camouflage, anti-aging properties, virulence and fungal pathogenesis. The above properties of melanin can be used in various application purposes which make the melanin expensive and great demand in the industry. To overcome the above situation, there is a necessity to find out the new biological sources for the maximum production of melanin. Numerous reports are available on fungal and bacterial melanin but only a few of them from actinomycetes. Based on the above view, the present investigation was aimed to purify the DOPA melanin from the novel marine actinomycetes Prauserella sp. (MAPPL 017A). Purified melanin was characterized using several state-of-the-art techniques and it’s showed the broadband absorbance, presence of carboxylic and indolic groups, high amount of carbon, nitrogen and low level of sulfur, presence of other metal ions such as calcium, sodium and magnesium with efficient thermal properties.

Unraveling the contributions to the neuromelanin-MRI contrast

The Locus Coeruleus (LC) and the Substantia Nigra (SN) are small brainstem nuclei that change with aging and may be involved in the development of various neurodegenerative and psychiatric diseases. Magnetization Transfer (MT) MRI has been shown to facilitate LC and the SN visualization, and the observed contrast is assumed to be related to neuromelanin accumulation. Imaging these nuclei may have predictive value for the progression of various diseases, but interpretation of previous studies is hindered by the fact that the precise biological source of the contrast remains unclear, though several hypotheses have been put forward. To inform clinical studies on the possible biological interpretation of the LC- and SN contrast, we examined an agar-based phantom containing samples of natural Sepia melanin and synthetic Cys-Dopa-Melanin and compared this to the in vivo human LC and SN. T1 and T2* maps, MT spectra and relaxation times of the phantom, the LC and the SN were measured, and a two-pool MT model was fitted. Additionally, Bloch simulations and a transient MT experiment were conducted to confirm the findings. Overall, our results indicate that Neuromelanin-MRI contrast in the LC likely results from a lower macromolecular fraction, thus facilitating interpretation of results in clinical populations. We further demonstrate that in older individuals T1 lengthening occurs in the LC.

Regulatory Effects of Nitric Oxide on Reproduction and Melanin Biosynthesis in Onion Pathogenic Fungus Stemphylium Eturmiunum

The formation of propagules (conidia and ascospores) is the critical stage for the transmission of the pathogenic fungus Stemphylium eturmiunum. However, how the development of these propagules is regulated remains to be fully understood. Here, we show that nitric oxide (NO) is necessary for the formation of conidia and pseudothecia in S. eturmiunum. Application of NO scavenger carboxy-CPTIO (cPTIO) or soluble guanylate cyclase (sGC) inhibitor NS2028 abolishes the formation of conidia and pseudothecia. In the culture of S. eturmiunum, supplement of NO-releasing compound sodium nitroprusside (SNP) results in an increased formation of conidia at 0.2 mmol/L, and pseudothecia at 2 mmol/L. SNP supplement also triggered increased biosynthesis of melanin, which can be inhibited partially upon the addition of either arbutin or tricyclazole, the specific inhibitors for 3,4-dihydroxyphenylalanine (DOPA) and dihydroxynaphthalene (DHN) synthetic pathway, respectively. Intriguingly, enhanced melanin biosynthesis triggered an increased formation of propagules; while its inhibition impaired their formation. The SNP-induced increment in the formation of propagules can be also compromised upon supplement of cPTIO or NS-2028. RT-PCR analysis showed that SNP at 0.2 mmol/L promoted transcription of the genes encoding the conidiation co-regulators brlA, abA, and wetA, and inhibited at 2 mmol/L. In contrast, application of SNP at 2 mmol/L increased transcription of the genes encoding mat1, and mat2, the genes related to sexual reproduction, and the transcription of two DNH melanin synthetic genes pks1 and pks2, and the key gene tyr for DOPA melanin biosynthesis. However, the increased transcription of these genes is down-regulated or blocked upon supplement of cPTIO or NS-2028. Thus, NO regulates asexual and sexual development, as well as melanin synthesis in S. eturmiunum possibly through NO-sGC-GMP signaling pathway.

Citric Acid Tunes the Formation of Antimicrobial Melanin-Like Nanostructures

Nature has provided a valuable source of inspiration for developing high performance multifunctional materials. Particularly, catechol-containing amino acid l-3,4-dihydroxyphenylalanine (l-DOPA) has aroused the interest to design hybrid multifunctional materials with superior adhesive ability. DOPA oxidative polymerization mediated by either melanogenic enzymes or an alkaline environment involving catechol intermolecular cross-linking, ultimately leads to melanin oligomers. Recently, relevant studies disclosed the ability of Ti-based nanostructures to tune melanin’s supramolecular structure during its formation, starting from melanogenic precursors, thus improving both antioxidant and antimicrobial properties. In this work, we propose a novel biomimetic approach to design hybrid DOPA melanin-like nanostructures through a hydrothermal synthesis opportunely modified by using citric acid to control hydrolysis and condensation reactions of titanium alkoxide precursors. UV-Vis and Electron paramagnetic resonance (EPR) spectroscopic evidences highlighted the key role of citrate–Ti(IV) and DOPA–Ti(IV) complexes in controlling DOPA polymerization, which specifically occurred during the hydrothermal step, mediating and tuning its conversion to melanin-like oligomers. Trasmission electron microscopy (TEM) images proved the efficacy of the proposed synthesis approach in tuning the formation of nanosized globular nanostructures, with high biocide performances. The obtained findings could provide strategic guidelines to set up biomimetic processes, exploiting the catechol-metal complex to obtain hybrid melanin-like nanosystems with optimized multifunctional behavior.