Roles of the P1, P2, and P3 Residues in Determining Inhibitory Specificity of Kallistatin toward Human Tissue Kallikrein

Chen, Vincent C.; Chao, Lee; Chao, Julie

doi:10.1074/jbc.m005605200

Cited by 42 publications

(30 citation statements)

References 43 publications

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“…However, SPN55 is cleaved between the putative P2 Tyr and P1 Met residues by the trypsin-like SAE, and the cleavage site of SPN48, which is targeted by the trypsin-like SPE, was determined to be between the putative P1Ј Glu and P2Ј Met residues. These unusual P1 specificities of the Arg-targeting SPs have also been observed in several mammalian serpins (12,27,28). For example, kallistatin, a serpin that inhibits tissue kallikrein, has a P1 Phe residue, despite the fact that kallikrein targets Arg-specific substrates (28).…”

Section: Discussionmentioning

confidence: 94%

“…These unusual P1 specificities of the Arg-targeting SPs have also been observed in several mammalian serpins (12,27,28). For example, kallistatin, a serpin that inhibits tissue kallikrein, has a P1 Phe residue, despite the fact that kallikrein targets Arg-specific substrates (28). Also, protein Z-dependent protease inhibitor, which has a P1 Tyr residue, is a specific inhibitor of membranebound factor Xa, an Arg-specific SP (12).…”

Section: Discussionmentioning

confidence: 97%

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Three Pairs of Protease-Serpin Complexes Cooperatively Regulate the Insect Innate Immune Responses

Jiang

Kim

Gong

et al. 2009

Journal of Biological Chemistry

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Serpins are known to be necessary for the regulation of several serine protease cascades. However, the mechanisms of how serpins regulate the innate immune responses of invertebrates are not well understood due to the uncertainty of the identity of the serine proteases targeted by the serpins. We recently reported the molecular activation mechanisms of three serine protease-mediated Toll and melanin synthesis cascades in a large beetle, Tenebrio molitor. Here, we purified three novel serpins (SPN40, SPN55, and SPN48) from the hemolymph of T. molitor. These serpins made specific serpin-serine protease pairs with three Toll cascade-activating serine proteases, such as modular serine protease, Spätzle-processing enzyme-activating enzyme, and Spätzle-processing enzyme and cooperatively blocked the Toll signaling cascade and ␤-1,3-glucanmediated melanin biosynthesis. Also, the levels of SPN40 and SPN55 were dramatically increased in vivo by the injection of a Toll ligand, processed Spätzle, into Tenebrio larvae. This increase in SPN40 and SPN55 levels indicates that these serpins function as inducible negative feedback inhibitors. Unexpectedly, SPN55 and SPN48 were cleaved at Tyr and Glu residues in reactive center loops, respectively, despite being targeted by trypsin-like Spätzle-processing enzyme-activating enzyme and Spätzle-processing enzyme. These cleavage patterns are also highly similar to those of unusual mammalian serpins involved in blood coagulation and blood pressure regulation, and they may contribute to highly specific and timely inactivation of detrimental serine proteases during innate immune responses. Taken together, these results demonstrate the specific regulatory evidences of innate immune responses by three novel serpins.

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Section: Discussionmentioning

confidence: 94%

Section: Discussionmentioning

confidence: 97%

Three Pairs of Protease-Serpin Complexes Cooperatively Regulate the Insect Innate Immune Responses

Jiang

Kim

Gong

et al. 2009

Journal of Biological Chemistry

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“…Kallistatin is mainly expressed in the liver but is also present in the heart, kidney, and blood vessel [21][22][23]. Kallistatin protein contains two structural elements: an active site and a heparin-binding domain [24][25][26]. The active site of kallistatin is necessary for complex formation with tissue kallikrein and thus inhibition of tissue kallikrein activity and bioavailability [13,27].…”

Section: Introductionmentioning

confidence: 99%

Kallistatin in Sepsis: Protective Actions and Potential Therapeutic Applications

Chao¹,

Li²,

Chao³

2017

Sepsis

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Sepsis is a systemic inlammatory response to infection, leading to multiorgan injury and mortality. Kallistatin is an endogenous protein expressed in the liver and tissues relevant to cardiovascular function. Kallistatin levels are markedly reduced in patients with sepsis and liver disease and in lipopolysaccharide (LPS)-induced septic mice. Kallistatin administration atenuates inlammation, multiorgan damage, and lethality in septic mice with LPS treatment, group A streptococcal, or polymicrobial infection. Importantly, kallistatin treatment not only prevents but also reverses organ injury and lethality in septic mice. Kallistatin decreases sepsis-induced inlammatory responses and tissue damage by modulating diferential signaling pathways, including: (1) stimulating endothelial nitric oxide (eNOS) and sirtuin 1 (SIRT) synthesis, and NO formation; (2) increasing suppressor of cytokine signaling-3 (SOCS3) expression; (3) antagonizing tumor necrosis factor-α (TNF-α) and high mobility group box 1 (HMGB1)-mediated oxidative stress and inlammatory gene expression; and (4) displaying bactericidal efects by stimulating superoxide formation. Therefore, kallistatin's multifactorial activities provide efective protection during septic shock in animal models. As kallistatin displays no apparent cytotoxicity, kallistatin therapy may provide a promising approach for the treatment of sepsis in humans.

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“…Additionally, amino acid sequence alignment of serpins shows that kallistatin has a unique insertion of 3-4 residues in the loop between the H helix and C2 sheet. A molecular model of kallistatin indicates that these extra residues may bulge the loop toward the reactive center loop (19). The basic amino acid residues K312:K313 in this loop are thus positioned in close vicinity to the reactive site (Fig.…”

mentioning

confidence: 99%

“…The high selectivity of kallistatin toward tissue kallikrein is attributed to its unique Phe-Phe pair at the P2-P1 residues in the reactive center (19). Kallistatin is also one of the heparinbinding serpins (18), which include antithrombin, protease nexin I, heparin cofactor II, plasminogen activator inhibitor, and protein C inhibitor (PCI).…”

mentioning

confidence: 99%

A Positively Charged Loop on the Surface of Kallistatin Functions to Enhance Tissue Kallikrein Inhibition by Acting as a Secondary Binding Site for Kallikrein

Chen

Chao

2000

Journal of Biological Chemistry

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Kallistatin is a serine proteinase inhibitor (serpin) that specifically inhibits tissue kallikrein. The inhibitory activity of kallistatin is abolished upon heparin binding. The loop between the H helix and C2 sheet of kallistatin containing clusters of basic amino acid residues has been identified as a heparin-binding site. In this study, we investigated the role of the basic residues in this region in tissue kallikrein inhibition. Kallistatin mutants containing double Ala substitutions for these basic residues displayed a 70 -80% reduction of association rate constants, indicating the importance of these basic residues in tissue kallikrein inhibition. A synthetic peptide derived from the sequence between the H helix and C2 sheet of kallistatin was shown to suppress the kallistatin-kallikrein interaction through competition for tissue kallikrein binding. To further evaluate the function of this loop, we used ␣1-antitrypsin, a nonheparin-binding serpin and slow tissue kallikrein inhibitor as a scaffold to engineer kallikrein inhibitors. An ␣1-antitrypsin chimera harboring the P3-P2 residues and a sequence homologous to the positively charged region between the H helix and C2 sheet of kallistatin acquired heparin-suppressed inhibitory activity toward tissue kallikrein and exhibited an inhibitory activity 20-fold higher than that of the other chimera, which contained only kallistatin's P3-P2 sequence, and 2300-fold higher than that of wild-type ␣1-antitrypsin. The ␣1-antitrypsin chimera with inhibitory characteristics similar to those of kallistatin demonstrates that the loop between the H helix and C2 sheet of kallistatin is crucial in tissue kallikrein inhibition, and this functional loop can be used as a module to enhance the inhibitory activity of a serpin toward tissue kallikrein. In conclusion, our results indicate that a positively charged loop between the H helix and C2 sheet of a serpin can accelerate the association of a serpin with tissue kallikrein by acting as a secondary binding site.

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Roles of the P1, P2, and P3 Residues in Determining Inhibitory Specificity of Kallistatin toward Human Tissue Kallikrein

Cited by 42 publications

References 43 publications

Three Pairs of Protease-Serpin Complexes Cooperatively Regulate the Insect Innate Immune Responses

Three Pairs of Protease-Serpin Complexes Cooperatively Regulate the Insect Innate Immune Responses

Kallistatin in Sepsis: Protective Actions and Potential Therapeutic Applications

A Positively Charged Loop on the Surface of Kallistatin Functions to Enhance Tissue Kallikrein Inhibition by Acting as a Secondary Binding Site for Kallikrein

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