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Bacterial collagen‐binding domain targets undertwisted regions of collagen
Author(s) -
Philominathan Sagaya Theresa Leena,
Koide Takaki,
Matsushita Osamu,
Sakon Joshua
Publication year - 2012
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1002/pro.2145
Subject(s) - heteronuclear single quantum coherence spectroscopy , chemistry , binding site , triple helix , collagen helix , n terminus , titration , stereochemistry , collagenase , biophysics , two dimensional nuclear magnetic resonance spectroscopy , crystallography , biochemistry , peptide sequence , enzyme , biology , inorganic chemistry , gene
Clostridium histolyticum collagenase causes extensive degradation of collagen in connective tissue that results in gas gangrene. The C‐terminal collagen‐binding domain (CBD) of these enzymes is the minimal segment required to bind to a collagen fibril. CBD binds unidirectionally to the undertwisted C‐terminus of triple helical collagen. Here, we examine whether CBD could also target undertwisted regions even in the middle of the triple helix. Collageneous peptides with an additional undertwisted region were synthesized by introducing a Gly → Ala substitution [(POG) x POA(POG) y ] 3 , where x + y = 9 and x > 3). 1 H– 15 N heteronuclear single quantum coherence nuclear magnetic resonance (HSQC NMR) titration studies with 15 N‐labeled CBD demonstrated that the minicollagen binds to a 10 Å wide 25 Å long cleft. Six collagenous peptides each labeled with a nitroxide radical were then titrated with 15 N‐labeled CBD. CBD binds to either the Gly → Ala substitution site or to the C‐terminus of each minicollagen. Small‐angle X‐ray scattering measurements revealed that CBD prefers to bind the Gly → Ala site to the C‐terminus. The HSQC NMR spectra of 15 N‐labeled minicollagen and minicollagen with undertwisted regions were unaffected by the titration of unlabeled CBD. The results imply that CBD binds to the undertwisted region of the minicollagen but does not actively unwind the triple helix.