Novel Pharmacophores of Connexin43 Based on the “RXP” Series of Cx43-Binding Peptides

Abstract

Abstract-Gap junction pharmacology is a nascent field. Previous studies have identified molecules that enhance intercellular communication, and may offer potential for innovative antiarrhythmic therapy. However, their specific molecular target(s) and mechanism(s) of action remain unknown. Previously, we identified a 34-aa peptide (RXP-E) that binds the carboxyl terminal domain of Cx43 (Cx43CT) and prevents cardiac gap junction closure and action potential propagation block. These results supported the feasibility of a peptide-based pharmacology to Cx43, but the structure of the core active element in RXP-E, an essential step for pharmacological development, remained undefined. Here, we used a combination of molecular modeling, surface plasmon resonance, nuclear magnetic resonance and patch-clamp strategies to define, for the first time, a unique ensemble of pharmacophores that bind Cx43CT and prevent closure of Cx43 channels. Two particular molecules are best representatives of this family: a cyclized heptapeptide (called CyRP-71) and a linear octapeptide of sequence RRNYRRNY. These 2 small compounds offer the first structural platform for the design of Cx43-interacting gap junction openers. Moreover, the structure of these compounds offers an imprint of a region of Cx43CT that is fundamental to gap junction channel function.

Glycogen synthase kinase 3β transfers cytoprotective signaling through connexin 43 onto mitochondrial ATP-sensitive K + channels

“…To facilitate rapid biodistribution and in vivo transduction into cardiomyocytes, the peptide was conjugated to TAT 47-57 as an intracellular carrier (19,20). TAT 47-57 -conjugated RRPPYN served as a control (13,19). Under normoxic conditions, RRNYRRNY had no effect on the viability of isolated cardiomyocytes compared with the control peptide RRPPYN or no peptide (Fig.…”

“…Although some compounds have been described that stimulate Cx43 and increase gap junction coupling, they also exert diverse Cx43-unrelated actions and/or their molecular targets remain elusive (26,27). Therefore, we chose RRNYRRNY, an RXP-related peptide that binds to the Cx43 C terminus (13,28). In mitoplast-attached recordings, RRNYRRNY significantly activated mitoK ATP channels in a Cx43-dependent manner.…”

“…In intact cells and the whole heart, Cx43 C-terminal binding peptides, such as RRNYRRNY, will also target sarcolemmal gap junctions and hemichannels (13,28). The precise role of gap junctions and Cx43 hemichannels in ischemia/reperfusion injury is unresolved, and it is unclear whether holding gap junctions …”

Pharmacological modulation of connexin‐formed channels in cardiac pathophysiology

“…Cyclo-Arg-Arg-Pro-Pro-Tyr-Arg-Gln More stable variant of RXP-E that prevents acidification-induced uncoupling (Verma et al, 2009) …”

“…However, when the cardiomyocytes were exposed to acidic conditions CTP-RXP-E preserved propagation of the action potential (Lewandowski et al, 2008). Furthermore, the core active structures (a linear octapeptide RRNYRRNY) were studied to identify the smallest Cx43 carboxyterminal domain binding molecule that was still active to regulate gap junctional communication (Verma et al, 2009). CyRPs were the first identified cyclic molecules that were able to bind Cx43 to stabilize the open state of the gap junction channels present at the plasma membrane.…”