Processing of Pro–Brain Natriuretic Peptide Is Suppressed by O-Glycosylation in the Region Close to the Cleavage Site

Abstract

Background: Processing of the brain natriuretic peptide (BNP) precursor, proBNP, is a convertase-dependent reaction that produces 2 molecules—the active BNP hormone and the N-terminal part of proBNP (NT-proBNP). Although proBNP was first described more than 15 years ago, very little is known about the cellular mechanism of its processing. The study of proBNP processing mechanisms is important, because processing impairments could be associated with the development of heart failure (HF). Methods: The biochemical properties of recombinant proBNP and NT-proBNP and the same molecules derived from the blood of HF patients were analyzed by gel-filtration chromatography, site-directed mutagenesis, and different immunochemical methods with a panel of monoclonal antibodies (MAbs). Results: Part of the proBNP molecule (amino acid residues 61–76) located near the cleavage site was inaccessible to specific MAbs because of the presence of O-glycans, whereas the same region in NT-proBNP was completely accessible. We demonstrated that a convertase (furin) could effectively cleave deglycosylated (but not intact) proBNP. Of several mutant proBNP forms produced in a HEK 293 cell line, only the T71A variant was effectively processed in the cell. Conclusions: Only proBNP that was not glycosylated in the region of the cleavage site could effectively be processed into BNP and NT-proBNP. Site-directed mutagenesis enabled us to ascertain the unique suppressing role of T71-bound O-glycan in proBNP processing.

Mechanism of FGF23 processing in fibrous dysplasia

“…There is precedent for similar coordinated O-glycosylation/furin protein processing in the work of Smenov and colleagues, in which they demonstrated that furin degradation of pro brain naturetic peptide is inhibited by O-glycosylation in HEK293 cells. (29) We tested this hypothesis in primary cultures of WT and mutant human BMSCs and found that there was increased cAMP in mutant BMSCs ( Fig. 2A) and a coordinated decrease in ppGalNAcT3 activity and increased furin activity (Fig.…”

Natriuretic Peptides in the Regulation of Cardiovascular Physiology and Metabolic Events

“…Cenderitide has also been shown to be more resistant to degradation by neutral endopeptidase neprilysin compared to ANP, BNP, and CNP 109. ProBNP has been shown to have 7 O‐glycosylation sites in the NT‐proBNP region and furin is effective in cleaving deglycosylated, but not intact proBNP 110, 111. A subsequent study showed that O‐glycosylation of T71 residue in proBNP attenuates its processing into active BNP 112.…”

Natriuretic Peptides and Analytical Barriers

“…ProBNP, however, is glycosylated both in the central region and in the region located close to the cleavage site, specifically at amino acid residues 63-76. This region can be blocked to sitespecific antibodies because of steric impediment due to glycosylation (20 ). The degree of glycosylation of both proBNP and NT-proBNP is highly individual.…”