Saxiphilin, a Saxitoxin-binding Protein with Two Thyroglobulin Type 1 Domains, Is an Inhibitor of Papain-like Cysteine Proteinases

• Published in: The Journal of biological chemistry 2000-05, Vol.275 (20), p.15572-15577
• Main Authors: Lenarčič, Brigita, Krishnan, Gomathi, Borukhovich, Renata, Ruck, Brian, Turk, Vito, Moczydlowski, Edward
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Amphibian Proteins; Animals; Carrier Proteins - chemistry; Carrier Proteins - pharmacology; Cathepsin B - antagonists & inhibitors; Cathepsin L; Cathepsins - antagonists & inhibitors; Cysteine Endopeptidases; cysteine proteinase; Cysteine Proteinase Inhibitors - chemistry; Cysteine Proteinase Inhibitors - pharmacology; Endopeptidases; Humans; Kinetics; Molecular Sequence Data; papain; Papain - antagonists & inhibitors; Rana catesbeiana; Recombinant Proteins - chemistry; Recombinant Proteins - pharmacology; Repetitive Sequences, Amino Acid; saxiphilin; saxitoxin; Sequence Alignment; Sequence Homology, Amino Acid; Thyroglobulin - chemistry
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M001406200

The type 1 domain of thyroglobulin is a protein module (Thyr-1) that occurs in a variety of secreted and membrane proteins. Several examples of Thyr-1 modules have been previously identified as inhibitors of the papain family of cysteine proteinases. Saxiphilin is a neurotoxin-binding protein from bullfrog and a homolog of transferrin with a pair of such Thyr-1 modules located in the N-lobe. Saxiphilin is now characterized as a potent inhibitor of three cysteine proteinases as follows: papain, human cathepsin B, and cathepsin L. The stoichiometry of enzyme inhibition reveals that both Thyr-1 domains of saxiphilin inhibit papain (apparentKi = 1.72 nm), but only one of these domains inhibits cathepsin B (Ki = 1.67 nm) and cathepsin L (Ki = 0.02 nm). Physical association of saxiphilin and papain blocked from turnover at the active-site cysteine residue can be detected by cross-linking with glutaraldehyde. The rate of association of saxiphilin and cathepsin B is strongly pH-dependent with an optimum at pH 5.2, reflecting control by at least two H+-titratable groups. These results further demonstrate that various Thyr-1 domains are selective inhibitors of cysteine proteinases with utility in the study of protein interactions and degradation.