The Corn Inhibitor of Activated Hageman Factor: Purification and Properties of Two Recombinant Forms of the Protein

Hazegh‐Azam, Maryam; Kim, Seung-Sup; Masoud, Sameer; Andersson, Laura A.; White, Frank F.; Johnson, Lowell B.; Muthukrishnan, S.; Reeck, Gerald R.

doi:10.1006/prep.1998.0882

Cited by 14 publications

(11 citation statements)

References 23 publications

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“…CHFI-2345 lacks 11 amino acids and the first disulfide bond at the N terminus, and Cys 55 is replaced by Asp. This mutant is identical to a previously described shortened form of CHFI (49) except for the replacement of Cys 55 . Mutant CHFI-1234 lacks 24 amino acids at the C terminus and the fifth disulfide bond, with unpaired Cys 57 replaced by Asp.…”

Section: Resultsmentioning

confidence: 72%

“…Previous attempts to obtain recombinant CHFI resulted in the preparation of nonfunctional protein because of inclusion body formation during expression in E. coli (49, 50). Because CHFI has five disulfide bonds (29), correct folding of the molecule requires an oxidative environment (51).…”

Section: Resultsmentioning

confidence: 99%

“…However, the E. coli cytoplasm is slightly reducing (52). Ordinary E. coli strains, such as BL21 DE3 (49, 50), that were previously used for CHFI expression do not provide optimal conditions for disulfide bond formation because of the functions of the antioxidant glutathione and thioredoxin systems (52). Based on these observations, the E. coli Rosetta-Gami 2 DE3 strain with mutations in the glutathione and thioredoxin systems (53) was chosen as a host for the production of CHFI and its mutants.…”

Section: Resultsmentioning

confidence: 99%

“…The elimination of the first and the fifth disulfide bonds does not affect the folding of the contact region of the truncated protein with FXIIa. Considering the conformation-based concept of the structure-activity investigation (55) and the observation (49) that a mutant that lacks 11 N-terminal amino acids and has a Cys 55 substitution is indistinguishable from full-size recombinant CHFI, we investigated the inhibitory activity of four truncated CHFI mutants in comparison with the wild-type and full-size recombinant CHFI (CHFI-12345). The shortened mutants CHFI-1234 ( K i = 1.0 ± 0.3 n m for FXIIa) and CHFI-2345 ( K i = 1.0 ± 0.2 n m ) retain the ability to inhibit FXIIa to the same extent as wild-type CHFI ( K i = 1.0 ± 0.1 n m ); this finding is consistent with previously reported results (49).…”

Section: Discussionmentioning

confidence: 99%

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Interactions Outside the Proteinase-binding Loop Contribute Significantly to the Inhibition of Activated Coagulation Factor XII by Its Canonical Inhibitor from Corn

Корнеева

Trubetskov

Korshunova³

et al. 2014

Journal of Biological Chemistry

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Background: Canonical inhibitors interact with cognate proteases through a protease-binding loop.Results: Protease-binding loop of corn Hageman factor inhibitor (CHFI) does not inhibit its cognate enzyme-activated coagulation factor XII (FXIIa).Conclusion: Nonloop regions of CHFI are required for specific inhibition of FXIIa.Significance: CHFI is the first canonical inhibitor whose protease-binding loop is not sufficient for cognate enzyme inhibition.

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Section: Resultsmentioning

confidence: 72%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 2 more Smart Citations

Interactions Outside the Proteinase-binding Loop Contribute Significantly to the Inhibition of Activated Coagulation Factor XII by Its Canonical Inhibitor from Corn

Корнеева

Trubetskov

Korshunova³

et al. 2014

Journal of Biological Chemistry

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“…CTI inhibits β XIIa [53] and has relative specificity for this form of XIIa [54]. The β form of XIIa does not bind to surfaces and has little ability to activate XI, although it does activate Pre-Kallikrein (which in turn amplifies XIIa production).…”

Section: Discussionmentioning

confidence: 99%

Systems Biology of Coagulation Initiation: Kinetics of Thrombin Generation in Resting and Activated Human Blood

et al. 2010

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Blood function defines bleeding and clotting risks and dictates approaches for clinical intervention. Independent of adding exogenous tissue factor (TF), human blood treated in vitro with corn trypsin inhibitor (CTI, to block Factor XIIa) will generate thrombin after an initiation time (Ti) of 1 to 2 hours (depending on donor), while activation of platelets with the GPVI-activator convulxin reduces Ti to ∼20 minutes. Since current kinetic models fail to generate thrombin in the absence of added TF, we implemented a Platelet-Plasma ODE model accounting for: the Hockin-Mann protease reaction network, thrombin-dependent display of platelet phosphatidylserine, VIIa function on activated platelets, XIIa and XIa generation and function, competitive thrombin substrates (fluorogenic detector and fibrinogen), and thrombin consumption during fibrin polymerization. The kinetic model consisting of 76 ordinary differential equations (76 species, 57 reactions, 105 kinetic parameters) predicted the clotting of resting and convulxin-activated human blood as well as predicted Ti of human blood under 50 different initial conditions that titrated increasing levels of TF, Xa, Va, XIa, IXa, and VIIa. Experiments with combined anti-XI and anti-XII antibodies prevented thrombin production, demonstrating that a leak of XIIa past saturating amounts of CTI (and not “blood-borne TF” alone) was responsible for in vitro initiation without added TF. Clotting was not blocked by antibodies used individually against TF, VII/VIIa, P-selectin, GPIb, protein disulfide isomerase, cathepsin G, nor blocked by the ribosome inhibitor puromycin, the Clk1 kinase inhibitor Tg003, or inhibited VIIa (VIIai). This is the first model to predict the observed behavior of CTI-treated human blood, either resting or stimulated with platelet activators. CTI-treated human blood will clot in vitro due to the combined activity of XIIa and XIa, a process enhanced by platelet activators and which proceeds in the absence of any evidence for kinetically significant blood borne tissue factor.

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A new enzyme-linked immunosorbent assay recognizing both rat and human activated coagulation Factor XII (FXIIa)

Papageorgiou

Yeo

Backx

et al. 2012

Journal of Immunological Methods

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The Corn Inhibitor of Activated Hageman Factor: Purification and Properties of Two Recombinant Forms of the Protein

Cited by 14 publications

References 23 publications

Interactions Outside the Proteinase-binding Loop Contribute Significantly to the Inhibition of Activated Coagulation Factor XII by Its Canonical Inhibitor from Corn

Interactions Outside the Proteinase-binding Loop Contribute Significantly to the Inhibition of Activated Coagulation Factor XII by Its Canonical Inhibitor from Corn

Systems Biology of Coagulation Initiation: Kinetics of Thrombin Generation in Resting and Activated Human Blood

A new enzyme-linked immunosorbent assay recognizing both rat and human activated coagulation Factor XII (FXIIa)

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