Exp Mol Med.  2014 Sep;46(9):e114. 10.1038/emm.2014.57.

An antibody reactive to the Gly63-Lys68 epitope of NT-proBNP exhibits O-glycosylation-independent binding

• Lee Y
• Kim H
• Chung J

• KMID: 2155379
• DOI: http://doi.org/10.1038/emm.2014.57

Abstract

The N-terminal fragment of prohormone brain natriuretic peptide (NT-proBNP) is a commonly used biomarker for the diagnosis of congestive heart failure, although its biological function is not well known. NT-proBNP exhibits heavy O-linked glycosylation, and it is quite difficult to develop an antibody that exhibits glycosylation-independent binding. We developed an antibody that binds to the recombinant NT-proBNP protein and its deglycosylated form with similar affinities in an enzyme immunoassay. The epitope was defined as Gly63-Lys68 based on mimetic peptide screening, site-directed mutagenesis and a competition assay with a peptide mimotope. The nearest O-glycosylation residues are Thr58 and Thr71; therefore, four amino acid residues intervene between the epitope and those residues in both directions. In conclusion, we report that an antibody reactive to Gly63-Lys68 of NT-proBNP exhibits O-glycosylation-independent binding.

MeSH Terms

Amino Acid Sequence
Animals
Antibodies/*immunology
Antigen-Antibody Reactions
Epitope Mapping
Epitopes/chemistry/genetics/*immunology
Glycosylation
HEK293 Cells
Heart Failure/immunology
Humans
Molecular Sequence Data
Mutagenesis, Site-Directed
Natriuretic Peptide, Brain/chemistry/genetics/*immunology
Peptide Fragments/chemistry/genetics/*immunology
Rabbits
Recombinant Fusion Proteins/chemistry/genetics/immunology
Antibodies
Epitopes
Natriuretic Peptide, Brain
Peptide Fragments
Recombinant Fusion Proteins