Refolded scFv antibody fragment against myoglobin shows rapid reaction kinetics

Cited 9 time in scopus
Metadata Downloads
Title
Refolded scFv antibody fragment against myoglobin shows rapid reaction kinetics
Author(s)
Hyeong Nam Song; J H Jang; Y W Kim; D H Kim; Sung Goo Park; Myung Kyu Lee; S H Paek; Eui-Jeon Woo
Bibliographic Citation
International Journal of Molecular Sciences, vol. 15, no. 12, pp. 23658-23671
Publication Year
2014
Abstract
Myoglobin is one of the early biomarkers for acute myocardial infarction. Recently, we have screened an antibody with unique rapid reaction kinetics toward human myoglobin antigen. Antibodies with rapid reaction kinetics are thought to be an early IgG form produced during early stage of in vivo immunization. We produced a recombinant scFv fragment for the premature antibody from Escherichia coli using refolding technology. The scFv gene was constructed by connection of the VH-VL sequence with a (Gly4Ser)3 linker. The scFv fragment without the pelB leader sequence was expressed at a high level, but the solubility was extremely low. A high concentration of 8 M urea was used for denaturation. The dilution refolding process in the presence of arginine and the redox reagents GSH and GSSH successfully produced a soluble scFv protein. The resultant refolded scFv protein showed association and dissociation values of 9.32 × 10 -4M -1·s -1and 6.29 × 10 -3 s -1, respectively, with an affinity value exceeding 107 M -1(kon/koff), maintaining the original rapid reaction kinetics of the premature antibody. The refolded scFv could provide a platform for protein engineering for the clinical application for diagnosis of heart disease and the development of a continuous biosensor.
Keyword
Acute myocardial infarctionMyoglobinPremature antibodySingle-chain variable fragment (scFv)
ISSN
1422-0067
Publisher
MDPI
DOI
http://dx.doi.org/10.3390/ijms151223658
Type
Article
Appears in Collections:
Division of Biomedical Research > Disease Target Structure Research Center > 1. Journal Articles
Files in This Item:

Items in OpenAccess@KRIBB are protected by copyright, with all rights reserved, unless otherwise indicated.