Glycosylation of PCSK9 Impairs its Interaction with LDLR But Assures its Stability

Proprotein convertase subtilisin/kexin type 9 (PCSK9) increases plasma LDL levels due to the degradation of the LDL receptor (LDLR). Anti PCSK9 antibody treatment leads to up to 70% reduction in plasma LDL levels. However, for poorly understood reasons, the treatment has no effect in some patients. Glycosylation is one of several PCSK9 post-translational modifications, with inconclusive effects on the protein’s stability and function. We tested the hypothesis that glycosylation increases PCSK9 plasma stability by reducing its interaction with LDLR and promoting its secretion from hepatocytes. We studied the implications of glycosylation on PCSK9 kinetics and function in vitro and with a patient cohort treated with Anti-PCSK9 antibodies. We analyzed the PCSK9 protein sequence to identify potential sites for N and O-glycosylation. We confirmed the N-glycosylation sequence and found 12 potential sites for O-glycosylation. To assess the effect of glycosylation on extracellular stability and function, we incubated for 24 hours: PCSK9-overexpressing HEPG2 cells with two glycosylation inhibitors and a human recombinant PCSK9 with deglycosylating enzymes. Following the ablation of the N-glycosylation, the levels of extracellular PCSK9 decreased while its affinity for LDLR increased. These results were also observed for O-glycosylation but with a lesser effect. Finally, we show that anti PCSK9 antibody treated patients, have a preponderance of circulating glycosylated PCSK9. In conclusion, N-glycosylation enhances PCSK9 stability; however, it decreases its binding to LDLR. Circulating PCSK9 glycosylation patterns and quantity might explain the differential response of patients to anti-PCSK9 treatment.