Epitope tagging for tracking elastin-like polypeptides.
Elastin-like polypeptides (ELPs) are a class of biocompatible, non-immunogenic and crosslinkable biomaterials that offer promise for use as an injectable scaffold for cartilage repair. In this study, an oligohistidine (His(6)) epitope tag was incorporated at the N-terminus of an ELP using recombinant DNA techniques to permit tracking without compromising on material biocompatibility. His(6)-tagged ELPs were successfully detected by Western blot analysis and quantified by ELISAs following digestion with trypsin. The mass of His(6) tagged ELP fragments freed from a crosslinked ELP hydrogel after digestion with trypsin correlated highly with hydrogel weight loss, providing evidence of the tag's capability to enable tracking of enzymatic degradation of the ELP hydrogel. The His(6) tag also facilitated recognition of crosslinked ELPs from background staining of articular cartilage. These results suggest that the His(6) epitope tag has the potential to track ELP scaffold loss independently of newly formed tissue mass for evaluating matrix remodeling in vivo.
Ong
Shin R
SR
Department of Biomedical Engineering, Duke University, 136 Hudson Hall, Box 90821, Durham, NC 27708, USA.
Trabbic-Carlson
Kimberly A
KA
Nettles
Dana L
DL
Lim
Dong W
DW
Chilkoti
Ashutosh
A
Setton
Lori A
LA
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001672:Biocompatible Materials / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D001673:Biodegradation, Environmental; D002358:Cartilage, Articular / Q000737:chemistry* / Q000276:immunology / Q000293:injuries; D000939:Epitopes / Q000032:analysis; D020100:Hydrogels / Q000032:analysis* / Q000378:metabolism; D008969:Molecular Sequence Data; D010455:Peptides / Q000032:analysis* / Q000737:chemistry / Q000235:genetics; D015202:Protein Engineering; D013552:Swine; D014357:Trypsin / Q000737:chemistry / Q000378:metabolism; D000097825:Elastin-Like Polypeptides
D001672:Biocompatible Materials; D000939:Epitopes; D020100:Hydrogels; D010455:Peptides; D014357:Trypsin; C509318:His6-tagged elastin-like peptide; D000097825:Elastin-Like Polypeptides