Predictions Suggesting a Participation of β-Sheet Configuration in the M2 Domain of the P2X7 Receptor: A Novel Conformation?

Abstract

Scanning experiments have shown that the putative TM2 domain of the P2X(7) receptor (P2X(7)R) lines the ionic pore. However, none has identified an alpha-helix structure, the paradigmatic secondary structure of ion channels in mammalian cells. In addition, some researchers have suggested a beta-sheet conformation in the TM2 domain of P2X(2). These data led us to investigate a new architecture within the P2X receptor family. P2X(7)R is considered an intriguing receptor because its activation induces nonselective large pore formation, in contrast to the majority of other ionic channel proteins in mammals. This receptor has two states: a low-conductance channel (approximately 10 pS) and a large pore (> 400 pS). To our knowledge, one fundamental question remains unanswered: Are the P2X(7)R channel and the pore itself the same entity or are they different structures? There are no structural data to help solve this question. Thus, we investigated the hydrophobic M2 domain with the aim of predicting the fitted position and the secondary structure of the TM2 segment from human P2X(7)R (hP2X(7)R). We provide evidence for a beta-sheet conformation, using bioinformatics algorithms and molecular-dynamics simulation in conjunction with circular dichroism in different environments and Fourier transform infrared spectroscopy. In summary, our study suggests the possibility that a segment composed of residues from part of the M2 domain and part of the putative TM2 segment of P2X(7)R is partially folded in a beta-sheet conformation, and may play an important role in channel/pore formation associated with P2X(7)R activation. It is important to note that most nonselective large pores have a transmembrane beta-sheet conformation. Thus, this study may lead to a paradigmatic change in the P2X(7)R field and/or raise new questions about this issue.

P2X7 receptor-mediated calcium dynamics in HEK293 cells: experimental characterization and modelling approach

“…In particular, the results reported in [18] suggest the coexistence of two functions (channel and large pore) in a single structure. Moreover, even if chemical crosslinking and blue native polyacrylamide gel electrophoresis (BN-PAGE) analysis of recombinant receptors have revealed a trimeric structure of P2X ion channels, which has been confirmed by atomic force microscopy and by crystallography, the P2X7 functional structure is still far to be determined [18,19]. Recently, we proposed a possible functional and structural model of the P2X7R, in which the receptor could be composed of two conductive pathways: the first with the ATPbinding sites and is characterized by sensitivity to extracellular Mg 2+ and selective permeability to Ca 2+ (P2X7R-A); the second one permeable to cations and with activation and divalent cation sensitivity controlled by the first pathway (P2X7R-B) [20].…”

“…This has been linked to apoptosis [14][15][16] and many hypotheses have been proposed to explain this behaviour but the question is still open [17]. In particular, the results reported in [18] suggest the coexistence of two functions (channel and large pore) in a single structure. Moreover, even if chemical crosslinking and blue native polyacrylamide gel electrophoresis (BN-PAGE) analysis of recombinant receptors have revealed a trimeric structure of P2X ion channels, which has been confirmed by atomic force microscopy and by crystallography, the P2X7 functional structure is still far to be determined [18,19].…”

P2X purinergic receptor ligands: recently patented compounds

“…P2X7R differs from the other P2X receptors by its long, 240-amino acid carboxy-terminus tail, which is commonly believed to be necessary for its cell permeabilizing property [14]. Until only very recently, there was no crystal structure or suitable homology model available to assist structure-based drug design for P2X receptor families [15][16][17]. Most novel compounds for these receptors emerge as a result of high-throughput screening from pharmaceutical companies.…”