The ability of enzymes to recognize and process structurally diverse substrates is fundamental to metabolic flexibility and biological regulation. In melanin biosynthesis, human tyrosinase (Tyr) catalyzes the oxidation of several chemically distinct intermediates, including L-tyrosine, L-DOPA, DHICA, and DHI. Although its catalytic chemistry is well established, the structural basis of substrate selectivity and how it is altered by disease-associated mutations remains unclear. Using molecular docking and molecular dynamics simulations, we mapped the Tyr active site and identified 23 evolutionarily conserved residues that mediate multi-substrate recognition and binding. Across all substrates, binding induces coordinated conformational responses, particularly within an anchoring region (334-347) that provides electrostatic and hydrophobic steering, and a flexible gating loop (374-386) that modulates access and stabilizes bound intermediates. The OCA1B-associated P406L mutation, although distant from the catalytic core, disrupts long-range dynamic coupling and impairs loop flexibility, while 25 ClinVar-listed genetic variants at substrate-interacting residues weaken active-site organization, underscoring the sensitivity of Tyr's dynamic network to perturbation. Integrating these findings, we propose an ordered multi-substrate binding mechanism in which substrates are first guided by the anchoring region, then aligned by the universal triad, and finally refined through loop-mediated, substrate-specific contacts. Our work suggests a dynamic framework that could be useful for understanding human tyrosinase catalysis, genetic mutation impact, and future engineering strategies.

Oculocutaneous albinism type 1 (OCA1) caused by pathogenic variants of the TYR gene is an autosomal recessive disorder of pigmentation characterized by reduced biosynthesis of melanin pigment in skin, hair, and eyes. We had the opportunity to examine five East Cameroon Baka rainforest hunter-gatherers (historically called "Pygmies") with albinism and belonging to three different families. Screening of known albinism genes revealed a homozygous missense variant in the TYR gene, NM_000372.5: c.1109T>C; p.Met370Thr. In addition, one patient was also hemizygous for a variant in GPR143, the gene involved in ocular albinism (OA1). Another patient was also heterozygous for the common African and Afro-American 2.7-kb deletion in the OCA2 gene indicating admixture of one parent with neighboring Nzimé Bantu-speaking farmers. This is the first report of the occurrence of OCA1 in African rainforest hunter-gatherers.

Albino (Tyrc-2J/Tyrc-2J) C57BL/6J mice carry a mutation in the tyrosinase gene and are known to display alterations of photoreceptor synaptic ribbons. In the present study, we wanted to test whether similar alterations exist in oculocutaneous albinism type 1 (OCA1), a human disease that also results from mutations in the tyrosinase gene. In the present study, we assessed the morphology of a human OCA1 retina in comparison to control human retinas. We analyzed the retina of a 35-year-old OCA1 patient by immunolabeling at light and electron microscopic levels, conventional transmission electron microscopy, and by genomic DNA sequencing of the RIBEYE/CtBP2 gene in comparison to normal human controls. The morphological analyses revealed an overall surprisingly normal appearance of the retina, except for the presence of strikingly abnormal photoreceptor synaptic ribbons. Synaptic ribbons are presynaptic specializations of the continuously active retinal ribbon synapses and mainly consist of the RIBEYE protein. In the OCA1 patient, photoreceptor synaptic ribbons were very small and reduced to small fragments that were either still associated with the active zone transmitter release site or floating in the cytosol. The RIBEYE gene appeared to be unaltered in the OCA1 patient, except for some single nucleotide polymorphisms (SNPs) that were also present in controls. The OCA1 patients displayed similar defects of photoreceptor synaptic ribbons as previously observed in the albinotic mice with a defect in the tyrosinase gene. The observed alterations of synaptic ribbons are not due to mutations in the RIBEYE gene but are likely indirect consequences of the deficient melanin biosynthesis in the OCA1 patient.

Oculocutaneous albinism type 1A (OCA1A) is a rare recessive genetic condition caused by mutations in TYROSINASE (TYR) that results in pigmentation defects of the skin, hair and eyes. This study was performed to understand melanosome biogenesis and maturation defects in an OCA1A in vitro model using retinal pigment epithelium (RPE) derived from TYR knockout human induced pluripotent stem cells (iPSC). CRISPR-Cas9 was used to knockout the TYR gene in iPSC to generate an isogenic pair. A developmentally guided protocol was used to differentiate the isogenic iPSC pair towards RPE monolayer tissue. Monolayer organization, melanosome formation and maturation were studied using electron microscopy. Loss of TYR protein was studied using Western blot and immuno-fluorescence staining. RPE cellular morphology and junction integrity was studied using immunofluorescence staining and transepithelial resistance measurements. An isogenic pair comprising of untargeted control and TYR knockout iPSC were successfully differentiated towards RPE monolayer tissue with polygonal cell morphology. TYR knockout RPE exhibited significantly reduced TYR protein, increased presence of immature pre-melanosomes and a complete lack of mature melanosomes. We observed abnormal junctional localization of β-catenin staining pattern, as has been reported previously for albino mouse RPE- and OCA1A patient-derived RPE. Differentiation of TYR-deficient iPSC toward RPE displayed pigmentation defects and absence of mature melanosomes, whereas melanosome biogenesis was not affected, because pre-melanosomes were still observed. These observations were also similar to what was observed in OCA1A patient-derived RPE monolayer tissue, independently confirming the validity of these previous findings.