Interaction of Collagen-Like Peptide Models of Asymmetric Acetylcholinesterase with Glycosaminoglycans:  Spectroscopic Studies of Conformational Changes and Stability

Abstract

The effect of heparin on the conformation and stability of triple-helical peptide models of the collagen tail of asymmetric acetylcholinesterase expands our understanding of heparin interactions with proteins and presents an opportunity for clarifying the nature of binding of ligands to collagen triple-helix domains. Within the collagen tail of AChE, there are two consensus sequences for heparin binding of the form BBXB, surrounded by additional basic residues. Circular dichroism studies were used to determine the effect of the addition of increasing concentrations of heparin on triple-helical peptide models for the heparin binding domains, including peptides in which the basic residues within and surrounding the consensus sequence were replaced by alanine residues. The addition of heparin caused an increased triple-helix content with saturation properties for the peptide modeling the C-terminal site, while precipitation, with no increased helix content resulted from heparin addition to the peptide modeling the N-terminal site. The results suggest that the two binding sites with a similar triple-helical conformation have distinctive ways of interacting with heparin, which must relate to small differences in the consensus sequence (GRKGR vs GKRGK) and in the surrounding basic residues. Addition of heparin increased the thermal stability of all peptides containing the consensus sequence. Heparan sulfate produced conformational and stabilization effects similar to those of heparin, while chondroitin sulfate led to a cloudy solution, loss of circular dichroism signal, and a smaller increase in thermal stability. Thus, specificity in both the sequence of the triple helix and the type of glycosaminoglycan is required for this interaction.

Two Different Heparin-binding Domains in the Triple-helical Domain of ColQ, the Collagen Tail Subunit of Synaptic Acetylcholinesterase

“…In this context, a possible candidate is chondroitin sulfate. It has been suggested previously that chondroitin sulfate proteoglycans interact with asymmetric AChE (34,35), and we have observed previously a direct interaction between chondroitin sulfate and the peptides modeling the N-and C-terminal HBDs, whose characteristics also differed between the two sites (14).…”

“…The effect of the distant environment may also explain the fact that the wild type enzyme with both HBDs exhibits a greater affinity for heparin than each HBD independently. We have previously shown that triple-helical content and stability increase upon heparin binding to a HBD (14). Thus, such a perturbation induced by heparin binding to one HBD could propagate along the triple helix, altering the conformation of the other HBD and increasing its affinity for heparin.…”

“…We have characterized the structural properties of those putative heparin-binding domains (HBDs) by using molecular modeling and synthetic peptides (12)(13)(14). Here, we analyzed ColQ-heparin interactions by heparin affinity chromatography of the entire A 12 AChE, carrying different mutations in ColQ.…”

Structural Requirements for Heparin/Heparan Sulfate Binding to Type V Collagen

“…Furthermore, the homotrimer has a triple-helical conformation, whereas HepV does not, as demonstrated by circular dichroism, and that might contribute to their different requirements for sulfate group binding. It has been previously reported for the heparin binding domains of the acetylcholinesterase collagen tail that the addition of increased amounts of heparin can increase the triple helix content and the thermal stability of triple-helical peptide models (40,41). However, the addition of heparin and heparan sulfate at different concentrations failed to induce significant structural change in HepV even during an overnight incubation.…”

Pathogenic Factors in Vascular Dementia and Alzheimer’s Disease

“…A histidine in the catalytic subunit of ACHE accelerates its rate of cationic ligand binding [103]. The ACHE collagen tail has 2 consensus sequences that bind heparin and are surrounded by additional basic residues which enhance the heparinbinding affinity [104]. Anionic heparin inhibition of cation effects, of basic amino acid residues (histidine, arginine, lysine) and of HSPG, strongly suggests that heparin would ameliorate the harmful effects of ACHE.…”