Paper Citing NAMD - Abstract
Korneeva, Vera A.; Trubetskov, Mikhail M.; Korshunova, Alena V.; Lushchekina, Sofya V.; Kolyadko, Vladimir N.; Sergienko, Olga V.; Lunin, Vladimir G.; Panteleev, Mikhail A.; Ataullakhanov, Fazoil I.
Interactions Outside the Proteinase-binding Loop Contribute Significantly to the Inhibition of Activated Coagulation Factor XII by Its Canonical Inhibitor from Corn
JOURNAL OF BIOLOGICAL CHEMISTRY, 289:14109-14120, MAY 16 2014
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. Activated factor XII (FXIIa) is selectively inhibited by corn Hageman factor inhibitor (CHFI) among other plasma proteases. CHFI is considered a canonical serine protease inhibitor that interacts with FXIIa through its protease-binding loop. Here we examined whether the protease-binding loop alone is sufficient for the selective inhibition of serine proteases or whether other regions of a canonical inhibitor are involved. Six CHFI mutants lacking different N- and C-terminal portions were generated. CHFI-234, which lacks the first and fifth disulfide bonds and 11 and 19 amino acid residues at the N and C termini, respectively, exhibited no significant changes in FXIIa inhibition (K-i = 3.2 +/- 0.4 nm). CHFI-123, which lacks 34 amino acid residues at the C terminus and the fourth and fifth disulfide bridges, inhibited FXIIa with a K-i of 116 +/- 16 nm. To exclude interactions outside the FXIIa active site, a synthetic cyclic peptide was tested. The peptide contained residues 20-45 (Protein Data Bank code 1BEA), and a C29D substitution was included to avoid unwanted disulfide bond formation between unpaired cysteines. Surprisingly, the isolated protease-binding loop failed to inhibit FXIIa but retained partial inhibition of trypsin (K-i = 11.7 +/- 1.2 m) and activated factor XI (K-i = 94 +/- 11 m). Full-length CHFI inhibited trypsin with a K-i of 1.3 +/- 0.2 nm and activated factor XI with a K-i of 5.4 +/- 0.2 m. Our results suggest that the protease-binding loop is not sufficient for the interaction between FXIIa and CHFI; other regions of the inhibitor also contribute to specific inhibition.
