New Control Mechanism Discovered in Innate Immune System

Proteases are enzymes that cleave other proteins. Most often, proteases occur in cascade networks, where a particular event triggers a chain reaction in which several proteases cleave and thereby activate each other. Most well-known is probably the coagulation cascade, which causes clotting of our blood when a vessel is punctured.

Inter-α-inhibitor heavy chain 4 (ITIH4) is a liver-produced plasma protein belonging to the inter–α-inhibitor/ITIH family of proteins that consists of bikunin and six different heavy chain proteins. This protein family is also referred to as inter–α-trypsin inhibitor proteins since bikunin displays a weak inhibitory effect on proteases for which a biological role is yet to be defined.


Biomedical Scientists at Aarhus University (Aarhus, Denmark) and a colleague investigated which other proteins in the blood the so-called mannan-binding lectin–associated serine protease (MASP) proteases interact with the complement cascade. To characterize in detail how ITIH4 inhibits the MASP proteases, they isolated both free ITIH4 and ITIH4 bound to the MASP-1 protease. By the use of X-ray small-angle scattering and electron microscopy, these samples were studied. Liquid chromatography tandem-mass spectrometry was performed using a Q-Exactive plus mass spectrometer (ThermoFisher Scientific, Waltham, MA, USA).

The scientists showed that ITIH4 is cleaved by several human proteases within a protease-susceptible region, enabling ITIH4 to function as a protease inhibitor. This is exemplified by its inhibition of mannan-binding lectin–associated serine protease-1 (MASP-1), MASP-2, and plasma kallikrein, which are key proteases for intravascular host defense. Mechanistically, ITIH4 acts as bait that, upon cleavage, forms a non-covalent, inhibitory complex with the executing protease that depends on the ITIH4 von Willebrand factor A domain. ITIH4 inhibits the MASPs by sterically preventing larger protein substrates from accessing their active sites, which remain accessible and fully functional toward small substrates.

The authors concluded that the active sites of MASP within the non-covalent ITIH4-MASP complexes are catalytically competent, but downstream cleavage of C2 and C4 are inhibited by physically blocking access of the scissile bonds to the active sites. Such activity was demonstrated in human and murine serum. ITIH4 was found to be cleaved by various proteases within the protease-susceptible region (PSR), suggesting that ITIH4 is a broad-acting inhibitor that targets numerous proteases. The study was published on January 8, 2021 in the journal Science Advances.

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Aarhus University
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