Apolipoprotein E binds to and reduces serum levels of DNA-mimicking, pyrrolated proteins

Abstract

Lysine N-pyrrolation, converting lysine residues to Nϵ-pyrrole-l-lysine, is a recently discovered post-translational modification. This naturally occurring reaction confers electrochemical properties onto proteins that potentially produce an electrical mimic to DNA and result in specificity toward DNA-binding molecules such as anti-DNA autoantibodies. The discovery of this unique covalent protein modification provides a rationale for establishing the molecular mechanism and broad functional significance of the formation and regulation of Nϵ-pyrrole-l-lysine–containing proteins. In this study, we used microbeads coupled to pyrrolated or nonpyrrolated protein to screen for binding activities of human serum-resident nonimmunoglobin proteins to the pyrrolated proteins. This screen identified apolipoprotein E (apoE) as a protein that innately binds the DNA-mimicking proteins in serum. Using an array of biochemical assays, we observed that the pyrrolated proteins bind to the N-terminal domain of apoE and that oligomeric apoE binds these proteins better than does monomeric apoE. Employing surface plasmon resonance and confocal microscopy, we further observed that apoE deficiency leads to significant accumulation of pyrrolated serum albumin and is associated with an enhanced immune response. These results, along with the observation that apoE facilitates the binding of pyrrolated proteins to cells, suggest that apoE may contribute to the clearance of pyrrolated serum proteins. Our findings uncover apoE as a binding target of pyrrolated proteins, providing a key link connecting covalent protein modification, lipoprotein metabolism, and innate immunity.

Glycolaldehyde is an endogenous source of lysine N-pyrrolation

“…The lysine N-pyrrolation, converting lysine residues to pyrK, is associated with an increased negative charge due to neutralization of the lysine residues and with conferring electrochemical properties onto proteins that potentially produce an electrical mimic to DNA (2). Our recent study has revealed that the apoE deficiency leads to significant accumulation of pyrrolated serum albumin and is associated with an enhanced immune response (3). This finding strongly suggested that a physiologically-relevant molecule might be involved in the lysine N-pyrrolation.…”

“…In our previous studies, we showed that the immunization of BALB/c mice with the pyrrolated proteins accelerated the production of autoantibodies (2). In addition, the age-dependent increase in the IgG and IgM titers to the dsDNA and pyrrolated proteins were observed in the SLE-prone MRL-lpr mice (2) and in the apoE-deficient mice (3). In the present study, to evaluate the biological significance of GA as a source of lysine N-pyrrolation, we examined the antigenic property of the GA-modified proteins and observed that the immunization with GA-modified BSA induced robust IgG production to the pyrrolated proteins ( Fig.…”

“…Strikingly, the deficiency of apoE was associated with a significant increase in the IgG and IgM titers against GA-derived epitopes. The apoE-deficient mice exhibit evidence of the markedly increased covalent modification of lysine residues in proteins as assessed by the formation of a major lipid peroxidation-derived lysine adduct, N ε -(8-carboxyoctanyl)lysine, and autoantibodies to oxidized lipoproteins (3,24). The most likely scenario is that the deficiency of apoE accelerates the peroxidation of fatty acids, which may be associated with the accumulation of GA and GA-modified proteins, leading to the overproduction of the autoantibodies to the pyrrolated proteins.…”

“…We have recently identified apoE as a binding target of pyrK and pyrrolated proteins and have shown that the apoE deficiency leads to the significant accumulation of pyrrolated serum proteins and to an enhanced immune response (3). Hence, to determine if the GA-modified proteins could show an affinity with serum proteins, including apoE, normal mouse serum was incubated with the beads coupled to the native BSA and GA-modified BSA (GA-BSA).…”

“…On the other hand, Hirose et al (3) recently identified apolipoprotein E (apoE) as an innate pyrrole-binding protein in human serum, which promotes the uptake of pyrrolated proteins by macrophages. They have also shown that the deficiency of apoE leads to a significant elevation of the pyrrolated proteins (3). In addition, significantly elevated levels of serum pyrrolated proteins compared to the normal control individuals have also been demonstrated.…”