An engineered alpha1 integrin-binding collagenous sequence.
Collagen is an extracellular matrix structural component that can regulate cellular processes through its interaction with the integrins, α1β1, α2β1, α10β1, and α11β1. Collagen-like proteins have been identified in a number of bacterial species. Here, we used Scl2 from Streptococcus pyogenes serotype M28 strain MGAS6274 as a backbone for the introduction of discrete integrin-binding sequences. The introduced sequences GLPGER, GFPGER, or GFPGEN did not affect triple helix stability of the Scl (Streptococcal collagen-like) protein. Using ELISA and surface plasmon resonance, we determined that Scl2(GLPGER) and Scl2(GFPGER) bound to recombinant human α1 and α2 I-domains in a metal ion-dependent manner and without a requirement for hydroxyproline. We predicted a novel and selective integrin-binding sequence, GFPGEN, through the use of computer modeling and demonstrated that Scl2(GFPGEN) shows specificity toward the α1 I-domain and does not bind the α2 I-domain. Using C2C12 cells, we determined that intact integrins interact with the modified Scl2 proteins with the same selectivity as recombinant I-domains. These modified Scl2 proteins also acted as cell attachment substrates for fibroblast, endothelial, and smooth muscle cells. However, the modified Scl2 proteins were unable to aggregate platelets. These results indicate that Scl2 is a suitable backbone for the introduction of mammalian integrin-binding sequences, and these sequences may be manipulated to individually target α1β1 and α2β1.
Seo
Neungseon
N
Center for Infectious and Inflammatory Diseases, Institute of Biosciences and Technology, Texas A&M University System Health Science Center, Houston, Texas 77030, USA.
Russell
Brooke H
BH
Rivera
Jose J
JJ
Liang
Xiaowen
X
Xu
Xuejun
X
Afshar-Kharghan
Vahid
V
Höök
Magnus
M
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002448:Cell Adhesion; D002460:Cell Line; D003094:Collagen / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D006801:Humans; D039423:Integrin alpha1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D039421:Integrin alpha2 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000378:metabolism; D011485:Protein Binding; D015202:Protein Engineering / Q000379:methods*; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013297:Streptococcus pyogenes / Q000737:chemistry* / Q000235:genetics
D001426:Bacterial Proteins; D039423:Integrin alpha1; D039421:Integrin alpha2; D011994:Recombinant Proteins; C423453:Scl2 protein, Streptococcus; D003094:Collagen