Solution structure of the smallest cofactor-active fragment of thrombomodulin

• Published in: Nature structural & molecular biology Vol. 7; no. 3; pp. 200 - 204
• Main Authors: Komives, Elizabeth A, Wood, Matthew J, Sampoli Benitez, Benedetta A
• Format: Journal Article
• Language: English
• Published: United States Nature Publishing Group 01.03.2000
• Subjects: Acetylglucosamine - metabolism; Amino Acid Sequence; Animals; Binding Sites; Cattle; Disulfides - metabolism; Epidermal Growth Factor - chemistry; Glycosylation; Methionine - metabolism; Models, Molecular; Molecular Sequence Data; Molecular Weight; Motion; Nuclear Magnetic Resonance, Biomolecular; Peptide Fragments - chemistry; Peptide Fragments - metabolism; Pliability; Protein Structure, Tertiary; Solutions; Thrombin - metabolism; Thrombomodulin - chemistry; Thrombomodulin - metabolism
• ISSN: 1072-8368; 1545-9993; 2331-365X; 1545-9985
• DOI: 10.1038/73302

A glycosylated fragment of thrombomodulin containing two epidermal growth factor-like domains (TMEGF45) was analyzed by NMR. The 4th-domains structure of this two-domain fragment is similar to that of the individual domain previously determined. The 5th-domain, which has uncrossed disulfide bonds, is not as well determined in the two-domain fragment than the individual domain previously solved. The flexibility of the 5th-domain is consistent with low heteronuclear NOEs. In the individual 5th-domain, Met 388 was disordered, and key thrombin-binding residues formed a hydrophobic core. By contrast, in TMEGF45, Met 388 is in the 5th-domain core, positioned by Phe 376 from the 4th-domain. As a result, key thrombin-binding residues that were in the core of the individual domain are expelled. Upon thrombin binding, chemical shifts of two residues in the 4th-domain, the three interdomain linker residues, and nearly all of the 5th-domain are perturbed. Thus, TMEGF45 binds thrombin by an induced fit mechanism involving a flexible 5th-domain.