Publication Date

5-2022

Date of Final Oral Examination (Defense)

4-5-2022

Type of Culminating Activity

Dissertation

Degree Title

Doctor of Philosophy in Biomolecular Sciences

Department

Biology

Supervisory Committee Chair

Daniel Fologea, Ph.D.

Supervisory Committee Member

Denise Wingett, Ph.D.

Supervisory Committee Member

Matthew Ferguson, Ph.D.

Abstract

The defense work focused on deciphering novel functionalities of lysenin channels with respect to adjustment of their regulation and transport properties in response to environmental changes. Lysenin is a pore-forming toxin extracted from the earthworm E. fetida, which self-assembles into large pores in artificial and natural lipid membranes containing sphingomyelin. Prior investigations on their functionality identified strong regulation by voltage and ligands as fundamental traits of lysenin, similar to ion channels. In addition, stochastic sensing, controlled switching, and hysteretic conductance have been added to the list of intricate biophysical properties of lysenin channels as potential enablers of biotechnological and biomedical applications. Our work employed electrophysiological measurements of lysenin’s biophysical activity in various environmental conditions. The first important feature we discovered is the unusual selectivity of lysenin channels for monovalent anions and cations, which is uncommon for pore-forming toxins. In the same line, we exploited the ability of divalent cations to force the lysenin channels into a stable sub-conducting state and assessed how such conformational changes modulate their selectivity. Experimental and theoretical approaches indicated that sub-conductance is accompanied by a substantial loss of selectivity, leading to the dissipation of the electrical gradients. Next, our work focused on investigating the influence of Cu2+ ions on the functionality of lysenin channels. The major finding of these experiments is that Cu2+ addition influences the hysteretic conductance to such an extent that lysenin behaves like a memory molecule capable of preserving information about its immediate history in the absence of any applied voltage. Also, we identified that addition of Cu2+ restores the voltage-induced gating of lysenin channels reconstituted in neutral lipid membranes, known to suppress the voltage regulation. In conclusion, our results identified unusual biophysical properties of lysenin channels which may be further exploited not only for understanding novel biological activities of transmembrane transporters but also for potential use in biotechnology, bioengineering, and bioelectronics.

DOI

https://doi.org/10.18122/td.1918.boisestate

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