LDLRs and Genetic Causes of LDL Elevation Flashcards
Summarise the genetic and molecular structure of the LDLR.
These 860AA, 93kDa proteins that are formed of five domains, from the N-terminal extracellular domain to the C-terminal intracellular ones.
The gene is made up of 18 exons spanning 45kbp, producing a 5.3kbp transcript.
Describe domain I of the LDLR.
The N-terminal binding region has 7 40-AA Class A repeats (~50% identical), each with 6 cysteine residues that form disulphide bonds.
The original deletion studies showed that ApoB100 (hence LDL) binding requires only repeats 2-7, while only repeat 5 is needed for ApoE. However, this has now been contested, as it has been shown that deletion of repeats 1 and 2 confers familial hypercholesterolaemia.
Describe domain II of the LDLR.
This is a 400AA epidermal growth factor (EGF) precursor homology domain, named due to the ~30% homology with the EGF precursor gene.
This includes three “growth factor” repeats, A, B and C. A and B have binding activity, and are separated from C by a YWTD repeat region that forms a beta-propeller motif important for receptor recycling and releasing bound LDL in response to lower pH in the endosome.
Describe domain III of the LDLR.
This domain contains 2 N-linked and 18 O-linked oligosaccharides, which appear to have little function, as attested by its deletion, but may act as a spacer to ensure the binding region is beyond the extracellular matrix.
Describe domain IV of the LDLR.
This is the single helical transmembrane domain, comprising a chain of 22 hydrophobic amino acid residues.
Describe domain V of the LDLR.
The internal C-terminal domain (50 residues) contains a signal sequence (NPXY) essential for receptor-ligand (LDL) internalization. This interacts with clathrin, concentrating the LDLRs in the coated pits.
What triggers release of the LDL from the LDLR?
The release of the LDL when in the acidic endosome is thought to be mediated by the β-propellor region of the EGF precursor domain, which contains a large number of histidine residues that allow it to act as a sensor for the acidity. These have no net charge at physiological pH (7.4) but in the endosome (pH 5.3) they are protonated, leading to a conformational change in the domain.
A crystal structure of the extracellular domain at pH 5.3 showed the β-propellor domain interacting with the N-terminal Binding Region (repeat 4 and 5), hence it is thought that the change in conformation allows it to displace the LDL.
What is familial hypercholesterolaemia?
This is a family of genetic disorders characterised by incredibly high levels of plasma LDL, which leads to early onset CVD/CHD/atherosclerosis and xanthomas (deposition of LDL-derived cholesterol under the skin and in tendons).
In 90% of cases this is caused by mutations in the LDLR gene preventing proper uptake of LDLs.
What is the frequency and severity of FH?
Those heterozygous for the disorder tend to have 2-4 fold higher blood LDL levels, with 5-10 fold higher in homozygous individuals.
An autosomal dominant disorder, this has incidence (in Europe) of 1 in 500 (htz) and 1 in 106 (hmz); though recent next-gen sequencing studies suggest that it may in fact be as high as 1 in 200 and 160,000 respectively.
The high frequency is thought to be due to Alu repeats nearby and within the gene leading to instability through Alu-mediated recombination.
How are LDLR muations described and classified?
Around 90% of these are SNPs or small (
What are class 1 LDLR mutations?
No or negligible LDL-R synthesis in the endoplasmic reticulum (ER) – hence no surface expression.
What are class 2 LDLR mutations?
Defective transport to the Golgi body for glycosylation and sorting. About 50% of all LDL-R mutations are class 2. Usually, the receptor movement is delayed, rather than halted, and for a few mutants lacking the transmembrane domain the receptor is secreted. When this happens the mutation is labelled subclass 2B.
What is an example of a class 2A LDLR mutation?
In the Lebanese Mutation, the receptor is truncated after amino acid 659 - a nonsense mutation cuts off everything after the β-propeller region exposing two cysteine residues from glycan repeat C.
This prevents trafficking from the ER to the Golgi due to recruitment of ER ‘gatekeeper proteins’, resulting in degradation of the receptor protein without stimulation of an immune response.
The mutation is so known because it occurs in the Lebanon in higher frequencies, 1 in 100,000 rather than 1,000,000
What are class 3 LDLR mutations?
Mutations in the ligand binding repeats prevent binding of LDL. This can be tested for through the use of antibodies raised to wildtype LDLRs.
What are class 4 LDLR mutations?
Here binding of LDL is normal, but the mutant receptors are unable to internalise receptor-ligand complexes due to disruption of the 50-residues cytoplasmic domain (790-839). This prevents the receptors from congregating in clathrin coated pits or from internalising. Mutations that cause this generally occur early, as the first 22 amino acids are sufficient for rapid internalisation