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Identification of a human type XVII collagen fragment with high capacity for maintaining skin health.

ABSTRACT: Collagen XVII (COL17) is a transmembrane protein that mediates skin homeostasis. Due to expression of full length collagen was hard to achieve in microorganisms, arising the needs for selection of collagen fragments with desired functions for microbial biosynthesis. Here, COL17 fragments (27-33 amino acids) were extracted and replicated 16 times for recombinant expression in Escherichia coli. Five variants were soluble expressed, with the highest yield of 223 mg/L. The fusion tag was removed for biochemical and biophysical characterization. Circular dichroism results suggested one variant (sample-1707) with a triple-helix structure at >37 °C. Sample-1707 can assemble into nanofiber (width, 5.6 nm) and form hydrogel at 3 mg/mL. Sample-1707 was shown to induce blood clotting and promote osteoblast differentiation. Furthermore, sample-1707 exhibited high capacity to induce mouse hair follicle stem cells differentiation and osteoblast migration, demonstrating a high capacity to induce skin cell regeneration and promote wound healing. A strong hydrogel was prepared from a chitosan and sample-1707 complex with a swelling rate of >30 % higher than simply using chitosan. Fed-batch fermentation of sample-1707 with a 5-L bioreactor obtained a yield of 600 mg/L. These results support the large-scale production of sample-1707 as a biomaterial for use in the skin care industry.

SUBMITTER: Wang X

PROVIDER: S-EPMC11192991 | biostudies-literature | 2024 Dec

REPOSITORIES: biostudies-literature

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Identification of a human type XVII collagen fragment with high capacity for maintaining skin health.

Wang Xinglong X Yu Shuyao S Sun Ruoxi R Xu Kangjie K Wang Kun K Wang Ruiyan R Zhang Junli J Tao Wenwen W Yu Shangyang S Linghu Kai K Zhao Xinyi X Zhou Jingwen J

Synthetic and systems biotechnology 20240606 4

Collagen XVII (COL17) is a transmembrane protein that mediates skin homeostasis. Due to expression of full length collagen was hard to achieve in microorganisms, arising the needs for selection of collagen fragments with desired functions for microbial biosynthesis. Here, COL17 fragments (27-33 amino acids) were extracted and replicated 16 times for recombinant expression in <i>Escherichia coli</i>. Five variants were soluble expressed, with the highest yield of 223 mg/L. The fusion tag was remo ...[more]

PMID: 38911060

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Project description:Bullous pemphigoid (BP), the most common autoimmune blistering disease, is induced by autoantibodies to type XVII collagen (COL17). Previous studies demonstrated that COL17 harbors several epitopes targeted by autoreactive T and B cells and that the target epitopes change sequentially during the disease course. To elucidate the details of the humoral immune response to COL17, we used an active BP mouse model in which BP is induced by the adoptive transfer of spleen cells from wild-type mice immunized with human COL17-expressing skin grafting to immunodeficient COL17-humanized (Rag-2-/-, mouse Col17-/-, human COL17+) mice. By immunoblot analysis, antibodies to the NC16A domain and other extracellular domains (ECDs) of COL17 were detected earlier than antibodies to intracellular domains (ICDs) in the active BP model. Time course analysis by enzyme-linked immunosorbent assay demonstrated a delayed peak of antibodies to ICD epitopes in active BP model. The blockade of CD40-CD40 ligand interaction soon after the adoptive transfer suppressed the production of antibodies to the non-collagenous 16A (NC16A) domain but not to an ICD epitope, suggesting the sequential activation from T and B cells against the ECD epitopes including the NC16A domain to those against ICD epitopes in vivo. Both wild-type mice immunized with a fragment of the NC16A domain and the recipients of those spleen cells produced IgG antibodies to ICD and ECD epitopes, showing intramolecular epitope spreading from the NC16A domain to other epitopes of COL17. Furthermore, we found that a portion of the active BP model mice show intermolecular epitope spreading from human COL17 to murine BP230. The appearance of antibodies to ICD epitopes of COL17 or of antibodies to murine BP230 did not correlate with the skin changes in the mice, suggesting that those antibodies have low pathogenicity. These results suggest that the immune response to the ECD epitopes of COL17, especially to the NC16A domain, triggers intramolecular, and intermolecular epitope spreading to ICD epitopes of COL17 and to murine BP230. These novel findings provide insight into the mechanism of epitope spreading in organ-specific, antibody-mediated autoimmune disorders.

Dataset Information

Identification of a human type XVII collagen fragment with high capacity for maintaining skin health.

Publications

Identification of a human type XVII collagen fragment with high capacity for maintaining skin health.

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