Noncollagenous region of the streptococcal collagen-like protein is a trimerization domain that supports refolding of adjacent homologous and heterologous collagenous domains.
Proper folding of the (Gly-Xaa-Yaa)(n) sequence of animal collagens requires adjacent N- or C-terminal noncollagenous trimerization domains which often contain coiled-coil or beta sheet structure. Collagen-like proteins have been found recently in a number of bacteria, but little is known about their folding mechanism. The Scl2 collagen-like protein from Streptococcus pyogenes has an N-terminal globular domain, designated V(sp), adjacent to its triple-helix domain. The V(sp) domain is required for proper refolding of the Scl2 protein in vitro. Here, recombinant V(sp) domain alone is shown to form trimers with a significant alpha-helix content and to have a thermal stability of T(m) = 45 degrees C. Examination of a new construct shows that the V(sp) domain facilitates efficient in vitro refolding only when it is located N-terminal to the triple-helix domain but not when C-terminal to the triple-helix domain. Fusion of the V(sp) domain N-terminal to a heterologous (Gly-Xaa-Yaa)(n) sequence from Clostridium perfringens led to correct folding and refolding of this triple-helix, which was unable to fold into a triple-helical, soluble protein on its own. These results suggest that placement of a functional trimerization module adjacent to a heterologous Gly-Xaa-Yaa repeating sequence can lead to proper folding in some cases but also shows specificity in the relative location of the trimerization and triple-helix domains. This information about their modular nature can be used in the production of novel types of bacterial collagen for biomaterial applications.
Yu
Zhuoxin
Z
Department of Biochemistry, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, USA.
Mirochnitchenko
Oleg
O
Xu
Chunying
C
Yoshizumi
Ayumi
A
Brodsky
Barbara
B
Inouye
Masayori
M
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D002152:Calorimetry, Differential Scanning; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism; D003094:Collagen / Q000737:chemistry*; D019281:Dimerization; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013297:Streptococcus pyogenes / Q000378:metabolism*; D013329:Structure-Activity Relationship
D001426:Bacterial Proteins; D002352:Carrier Proteins; D011994:Recombinant Proteins; D003094:Collagen