Lysenin Channels as Sensors for Ions and Molecules

Abstract

Lysenin is a pore-forming protein extracted from the earthworm Eisenia fetida, which inserts large conductance pores in artificial and natural lipid membranes containing sphingomyelin. Its cytolytic and hemolytic activity is rather indicative of a pore-forming toxin; however, lysenin channels present intricate regulatory features manifested as a reduction in conductance upon exposure to multivalent ions. Lysenin pores also present a large unobstructed channel, which enables the translocation of analytes, such as short DNA and peptide molecules, driven by electrochemical gradients. These important features of lysenin channels provide opportunities for using them as sensors for a large variety of applications. In this respect, this literature review is focused on investigations aimed at the potential use of lysenin channels as analytical tools. The described explorations include interactions with multivalent inorganic and organic cations, analyses on the reversibility of such interactions, insights into the regulation mechanisms of lysenin channels, interactions with purines, stochastic sensing of peptides and DNA molecules, and evidence of molecular translocation. Lysenin channels present themselves as versatile sensing platforms that exploit either intrinsic regulatory features or the changes in ionic currents elicited when molecules thread the conducting pathway, which may be further developed into analytical tools of high specificity and sensitivity or exploited for other scientific biotechnological applications.

The Curious Case of Earthworms and COVID-19

“…It is also possible to tag lysenin, e.g., with polyhistidine or fluorescence tags [58], and this is often used to identify SM in membranes. Further, it is possible to module the action of the lysenin-induced channels by interactions with multivalent inorganic and organic cations [59].…”

The Ionic Selectivity of Lysenin Channels in Open and Sub-Conducting States

“…Interestingly, the voltage-gating feature requires anionic lipids in the host membrane and is suppressed when neutral lipids are used [13]. However, the gating induced by multivalent ions acting as ligands is preserved in both neutral and charged lipid membranes [14]. Previous investigations on lysenin suggest a weak cation selectivity [15,17], as inferred from measuring the membrane voltage achieved upon creating asymmetrical ionic conditions.…”

“…Another category of membrane transporters that shares similarities with ion channels are represented by pore-forming toxins (PFTs), which introduce conducting pathways in the host membrane, capable of sustaining high transport rates [10]. Some PFTs may also present regulation by physical and chemical stimuli [11][12][13][14], and selectivity [15][16][17][18][19]. Although the ionic selectivity of PFTs is generally far under that of ion channels, prior investigations provided valuable information on the nature of the selectivity filters and even allowed intentional selectivity modulation by chemical modifications [16].…”

“…The pore-forming mechanism comprises binding to sphingomyelin and deployment of beta-sheets through the host membrane [25,26]. Lysenin exhibits a high transport rate and is also regulated by voltage and ligands [12][13][14]. Interestingly, the voltage-gating feature requires anionic lipids in the host membrane and is suppressed when neutral lipids are used [13].…”

“…Although the physiological relevance of sub-conductance is poorly understood, adjustments of the ionic permeabilities in such sub-conducting states have been reported [27][28][29]. For a better understanding of how intermediate conductance states adjust the transport properties of protein pores, we exploited a unique feature of lysenin channels, which is the attainment of stable sub-conducting states in the presence of divalent ions (i.e., Ca 2+ ) [12,14]. In this line, our experimental work demonstrates that the selectivity of lysenin channels to monovalent ions is significantly diminished when they are subconducting, which leads to vanishing membrane voltages.…”