Discovery of anti-inflammatory physiological peptides that promote tissue repair by reinforcing epithelial barrier formation

上皮バリアを形成するペプチドJIPの発見 --JIPは上皮組織修復に貢献する--. 京都大学プレスリリース. 2021-11-18.Epithelial barriers that prevent dehydration and pathogen invasion are established by tight junctions (TJs), and their disruption leads to various inflammatory diseases and tissue destruction. However, a therapeutic strategy to overcome TJ disruption in diseases has not been established because of the lack of clinically applicable TJ-inducing molecules. Here, we found TJ-inducing peptides (JIPs) in mice and humans that corresponded to 35 to 42 residue peptides of the C terminus of alpha 1-antitrypsin (A1AT), an acute-phase anti-inflammatory protein. JIPs were inserted into the plasma membrane of epithelial cells, which promoted TJ formation by directly activating the heterotrimeric G protein G13. In a mouse intestinal epithelial injury model established by dextran sodium sulfate, mouse or human JIP administration restored TJ integrity and strongly prevented colitis. Our study has revealed TJ-inducing anti-inflammatory physiological peptides that play a critical role in tissue repair and proposes a previously unidentified therapeutic strategy for TJ-disrupted diseases

Novel Dextranase Catalyzing Cycloisomaltooligosaccharide Formation and Identification of Catalytic Amino Acids and Their Functions Using Chemical Rescue Approach

A novel endo-dextranase (PsDex) from Paenibacillus sp. was found to mainly produce isomaltotetraose and small amounts of cycloisomaltooligosaccharides (CIs) with a degree of polymerization=of 7-14 from dextran. The 1,696 amino-acid sequence belonging to the glycosyl hydrolase family (GH) 66 has a long insertion (632 residues; Thr451-Val1082), a portion of which shares identity (35% at Ala39-Ser1304 of PsDex) with Pro32-Ala755 of CI glucanotransferase (CITase), a GH 66 enzyme that catalyzes the formation of CIs from dextran. This homologous sequence (Val837-Met932 for PsDex and Tyr404-Tyr492 for CITase), similar to carbohydrate-binding module 35, was not found in other endo-dextranases (Dexs) devoid of CITase activity. These results support the classification of GH 66 enzymes into 3 types: (i) Dex showing only dextranolytic activity, (ii) Dex catalyzing hydrolysis with low cyclization activity, and (iii) CITase showing CI-forming activity with low dextranolytic activity. The fact that a C-terminal truncated enzyme (having Ala39-Ser1304) has 50% wild-type PsDex activity indicates that the C-terminal 392 residues are not involved in hydrolysis. GH 66 enzymes possess 4 conserved acidic residues (Asp189, Asp340, Glu412, and Asp1254 of PsDex) of catalytic candidates. Their amide-mutants decreased activity (1/1500- 1/40,000-time), and D1254N had 36% activity. A chemical rescue approach was applied to D189A, D340G, and E412Q using α-isomaltotetraosyl fluoride with NaN3. D340G or E412Q formed a β- or α-isomaltotetraosyl azide, respectively, strongly indicating that Asp340 and Glu412 are nucleophile and acid/base-catalyst, respectively. Interestingly, D189A synthesized small-sized dextran from α-isomaltotetraosyl fluoride in the presence of NaN3