Functional Characterization of p.(Arg160Gln) PCSK9 Variant Accidentally Found in a Hypercholesterolemic Subject

Area of Science:

• Genetics
• Biochemistry
• Molecular Biology

Background:

• Familial hypercholesterolaemia (FH) is an autosomal dominant disorder causing high LDL cholesterol due to impaired LDL clearance.
• Key genes in FH include LDL receptor (LDLr), Apolipoprotein B (APOB), and PCSK9.
• PCSK9 gain-of-function (GOF) variants increase LDLr degradation, while loss-of-function (LOF) variants reduce it, impacting FH diagnosis.

Purpose of the Study:

• To functionally characterize the p.(Arg160Gln) PCSK9 variant in a patient suspected of having FH.
• To determine the variant's impact on LDL receptor (LDLr) activity and PCSK9 binding affinity.
• To elucidate the molecular mechanism behind the variant's effect on LDLr clearance.

Main Methods:

• Assessed PCSK9 autocatalytic cleavage efficiency.
• Measured protein expression levels of the variant.
• Quantified the effect of p.(Arg160Gln) PCSK9 on LDLr activity and its binding affinity to LDLr.

Main Results:

• The p.(Arg160Gln) variant showed expression and processing similar to wild-type (WT) PCSK9.
• This variant exhibited reduced activity on LDLr, with 13% higher LDL internalisation compared to WT.
• The p.(Arg160Gln) PCSK9 variant displayed lower affinity for LDLr (EC50 8.6 ± 0.8) than WT (EC50 25.9 ± 0.7).

Conclusions:

• The p.(Arg160Gln) PCSK9 variant is a loss-of-function (LOF) variant.
• Its reduced activity stems from a displaced PCSK9 P' helix, destabilizing the PCSK9-LDLr complex.
• Functional characterization of PCSK9 variants is vital for supporting FH genetic diagnosis.