BIOCHEMICAL, STRUCTURAL, AND TRANSGLUTAMINASE SUBSTRATE PROPERTIES OF HUMAN LORICRIN, THE MAJOR EPIDERMAL CORNIFIED CELL-ENVELOPE PROTEIN

Loricrin is the major protein of the cornified cell envelope of terminally differentiated epidermal keratinocytes which functions as a physical barrier. In order to understand its properties and role in cornified cell envelope, we have expressed human loricrin from a full-length cDNA clone in bacteria and purified it to homogeneity. We have also isolated loricrin from newborn mouse epidermis. By circular dichroism and fluorescence spectroscopy, the in vivo mouse and bacterially expressed human loricrins possess no alpha or beta structure but have some organized structure in solution associated with their multiple tyrosines and can be reversibly denatured by either guanidine hydrochloride or temperature. The transglutaminase (TGase) 1, 2, and 3 enzymes expressed during epidermal differentiation utilized loricrin in vitro as a complete substrate, but the types of cross-linking were different. The TGase 3 reaction favored certain lysines and glutamines by forming mostly intrachain cross-links, whereas TGase 1 formed mostly large oligomeric complexes by interchain cross-links involving different lysines and glutamines. Together, the glutamines and lysines used in vitro are almost identical to those seen in vivo. The data support a hypothesis for the essential and complementary roles of both TGase 1 and TGase 3 in cross-linking of loricrin in vivo. Failure to cross-link loricrin by TGase 1 may explain the phenotype of lamellar ichthyosis, a disease caused by mutations in the TGase 1 gene

Sequence characterization and glycosylation sites identification of donkey milk lactoferrin by multiple enzyme digestions and mass spectrometry

Lactoferrin, a protein showing an array of biochemical properties, including immuno-modulation, iron-binding ability, as well as antioxidant, antibacterial and antiviral activities, but which may also represent a potential milk allergen, was isolated from donkey milk by ion exchange chromatography. The characterization of its primary structure, by means of enzymatic digestions, SPITC derivatization of tryptic digest, reversed-phase high performance liquid chromatography, electrospray and matrix-assisted laser desorption/ionization mass spectrometry, is reported. Our results allowed the almost complete characterization of donkey lactoferrin sequence, that, at least for the covered sequence, differs from the horse genomic deduced sequence (UniProtKB Acc. Nr. O77811) by five point substitutions located at positions 91 (Arg → His), 328 (Thr → Ile/Leu), 466 (Ala → Gly), 642 (Asn → Ser) and 668 (Ser → Ala). Analysis of the glycosylated protein showed that glycans in donkey lactoferrin are linked to the protein backbone via an amide bond to asparagine residues located at the positions 137, 281 and 476.</p

The cornified envelope: A model of cell death in the skin

The epidermis functions as a barrier against the environment by means of several layers of terminally differentiated, dead keratinocytes - the cornified layer, which forms the endpoint of epidermal differentiation and death. The cornified envelope replaces the plasma membrane of differentiating keratinocytes and consists of keratins that are enclosed within an insoluble amalgam of proteins, which are crosslinked by transglutaminases and surrounded by a lipid envelope. New insights into the molecular mechanisms and the physiological endpoints of cornification are increasing our understanding of the pathological defects of this unique form of programmed cell death, which is associated with barrier malfunctions and ichthyosis

Clinical correlations of recent developments in the pathogenesis of atopic dermatitis Dermatite atópica: implicações clínicas de avanços recentes na patogênese

Atopic dermatitis is a chronic inflammatory skin disease with a steadily increasing prevalence affecting 10-20 of infants and 1-3 of adults globally. It is often the first clinical manifestation of atopic disease preceding asthma and allergic rhinitis. Probably half of the children with atopic dermatitis develop some other form of atopic disease later in life. The pathogenesis involves a complex interplay of factors including genetic predisposition due to altered immune or skin barrier function, interactions with the environment such as food and allergen exposures, and infectious triggers of inflammation. In this review, we summarize the recent advances in understanding the contribution of different factors in the pathophysiology of atopic dermatitis and how insights provide new therapeutic potential for its treatment.<br>A dermatite atópica é uma doença cutânea inflamatória crônica cuja prevalência tem aumentado de forma constante, afetando 10-20% dos lactentes e 1-3% dos adultos em todo o mundo. Ela é freqüentemente a primeira manifestação clínica de doença atópica, precedendo a asma e a rinite alérgica. Provavelmente metade das crianças com dermatite atópica desenvolvem alguma outra forma de doença atópica em outras fases da vida. A patogenia envolve uma interação complexa entre fatores que incluem predisposição genética devido a uma função alterada da barreira cutânea ou imunológica, interações com o ambiente, tais como exposição a alimentos e alergenos, e desencadeadores infecciosos de inflamação. Nesta revisão, resumimos os avanços recentes na compreensão da contribuição de diferentes fatores à fisiopatologia da dermatite atópica e como os novos conhecimentos proporcionam novo potencial terapêutico