### abstract ###
The relationship between Apolipoprotein E and the aggregation processes of the amyloid peptide has been shown to be crucial for Alzheimer's disease.
The presence of the ApoE4 isoform is considered to be a contributing risk factor for AD.
However, the detailed molecular properties of ApoE4 interacting with the A peptide are unknown, although various mechanisms have been proposed to explain the physiological and pathological role of this relationship.
Here, computer simulations have been used to investigate the process of A interaction with the N-terminal domain of the human ApoE isoforms.
Molecular docking combined with molecular dynamics simulations have been undertaken to determine the A peptide binding sites and the relative stability of binding to each of the ApoE isoforms.
Our results show that from the several ApoE isoforms investigated, only ApoE4 presents a misfolded intermediate when bound to A. Moreover, the initial -helix used as the A peptide model structure also becomes unstructured due to the interaction with ApoE4.
These structural changes appear to be related to a rearrangement of the salt bridge network in ApoE4, for which we propose a model.
It seems plausible that ApoE4 in its partially unfolded state is incapable of performing the clearance of A, thereby promoting amyloid forming processes.
Hence, the proposed model can be used to identify potential drug binding sites in the ApoE4-A complex, where the interaction between the two molecules can be inhibited.
### introduction ###
Alzheimer's disease is one of the most common neurodegenerative diseases at the present time.
The disease is characterized by the formation of neurofibrillary tangles and plaques in the brain, leading to neuronal dysfunction, neuronal loss and finally death.
The main component of the plaques is the amyloid- peptide, a 39 43 amino acids long hydrophobic peptide generated by the cleavage of the amyloid precursor, which accumulates in the form of soluble and non-soluble aggregates.
The connection between Apolipoprotein E and AD is well established CITATION, CITATION.
Structurally, ApoE is a 299 residues protein with an N-terminal domain involved in binding to heparin, low density lipoprotein receptors and LDLR-related proteins CITATION, CITATION.
The C-terminal domain has been related to heparin and lipid binding CITATION, CITATION.
Three main isoforms have been described for human ApoE, i.e. ApoE2, ApoE3 and ApoE4.
The standard variant is ApoE3, while ApoE2 is defective for receptor binding, causing APOE 2/ 2 homozygotic individuals to have a higher predisposition to diseases related to high amounts of cholesterol and triglycerides CITATION, CITATION.
For ApoE4, the receptor binding affinity remains unaffected, but APOE 4/ 4 homozygotic individuals have higher risk for coronary heart disease and a significantly greater risk for developing AD.
CITATION, CITATION Around 80 percent of all AD cases are related to the genetic variance at the ApoE locus CITATION, CITATION .
The only difference between the ApoE isoforms is found in residues 112 and 158, where Cys112/Cys158 corresponds to ApoE2, Cys112/Arg158 to ApoE3, and Arg112/Arg158 to ApoE4.
The presence of cysteines at these positions confers oligomerization properties to ApoE.
Indeed, ApoE2 and ApoE3 are able to form disulfide-linked homo- and hetero-oligomers due to the presence of respectively two and one Cys residue.
ApoE4 lacks the possibility of strong disulfide linking; however, it is unclear whether weaker interactions could promote the oligomerization of ApoE4.
The Cys/Arg substitution in ApoE4 also has molecular impact in terms of intra-protein polar contacts: the orientation of Arg61 is different in ApoE4 compared to ApoE3; the orientation of Arg61 towards the C-terminal domain facilitates a salt bridge between Arg61 and Glu255.
The electrostatic interaction between Arg61 and Glu255 promotes an N- and C-domain interaction that packs the structure tighter, which seems crucial for the interaction of ApoE4 with triglyceride-rich lipoproteins.
The interaction between Arg61 and Glu255 is absent in ApoE3 leading to a more open structure and a preferential binding of phospholipid-rich high-density lipoproteins CITATION, CITATION.
Chemical and thermal denaturation experiments have shown that the most unstable structure belongs to ApoE4, which displays a partially unfolded intermediate containing some structure that may be related to the fact that ApoE4 enhances the deposition of A CITATION, CITATION .
Although different mechanisms have been proposed to explain the physiological and pathological relationship between ApoE and the A peptide, the details of the interaction between ApoE and A at a molecular level are unknown.
Such detailed knowledge is however important for the understanding of the pathological mechanisms of AD, and may also help to identify potential therapeutic target sites where the interaction between ApoE4 and A can be blocked.
In the present study we are using molecular docking simulations based on global minimum energy to investigate the interaction process of A with the N-terminal domain of the different ApoE isoforms in order to determine potential A peptide binding sites in ApoE.
In the next step, molecular dynamics calculations are undertaken to explore the conformational dynamics of ApoE under A interaction and evaluate the stability of each of the ApoE-A complexes.
From the analysis and the statistics of the electrostatic interactions of the three ApoE isoforms, we present a model explaining the role of the A -ApoE interaction and its relevance for AD.
