Structure of Native Protein C Inhibitor Provides Insight into Its Multiple Functions

• Published in: The Journal of biological chemistry Vol. 282; no. 18; pp. 13759 - 13768
• Main Authors: Li, Wei, Adams, Ty E., Kjellberg, Margareta, Stenflo, Johan, Huntington, James A.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 04.05.2007; American Society for Biochemistry and Molecular Biology
• Subjects: Basic Medicine; Binding Sites - physiology; Crystallography, X-Ray; Escherichia coli; Heparin - chemistry; Heparin - metabolism; Humans; Hydrophobic and Hydrophilic Interactions; Läkemedelskemi; Medical and Health Sciences; Medicin och hälsovetenskap; Medicinal Chemistry; Medicinska och farmaceutiska grundvetenskaper; Nuclear Magnetic Resonance, Biomolecular; Protein Binding - physiology; Protein C Inhibitor - chemistry; Protein C Inhibitor - genetics; Protein C Inhibitor - metabolism; Protein Structure, Secondary; Protein Structure, Tertiary - physiology; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Serine Proteinase Inhibitors - chemistry; Serine Proteinase Inhibitors - genetics; Serine Proteinase Inhibitors - metabolism; Structural Homology, Protein; Structure-Activity Relationship; Thyroxine-Binding Proteins - chemistry
• ISSN: 0021-9258; 1083-351X; 1083-351X
• DOI: 10.1074/jbc.M701074200

Protein C inhibitor (PCI) is a multifunctional serpin with wide ranging protease inhibitory functions, unique cofactor binding activities, and potential non-inhibitory functions akin to the hormone-transporting serpins. To gain insight into the molecular mechanisms utilized by PCI we developed a robust expression system in Escherichia coli and solved the crystal structure of PCI in its native state. The five monomers obtained from our two crystal forms provide an NMR-like ensemble revealing regions of inherent flexibility. The reactive center loop (RCL) of PCI is long and highly flexible with no evidence of hinge region incorporation into β-sheet A, as seen for other heparin-binding serpins. We adapted an extrinsic fluorescence method for determining dissociation constants for heparin and find that the N-terminal tail of PCI and residues adjacent to helix H are not involved in heparin binding. The minimal heparin length capable of tight binding to PCI was determined to be chains of eight monosaccharide units. A large hydrophobic pocket occupied by hydrophobic crystal contacts was found in an analogous position to the hormone-binding site in thyroxine-binding globulin. In conclusion, the data presented here provide important insights into the mechanisms by which PCI exercises its multiple inhibitory and non-inhibitory functions.