NMR Structure and Dynamics of the Starfish Three-Finger Protein Lystar5

Abstract Objective: The primary goal of this study was to determine the spatial structure and investigate the intramolecular dynamics of Lystar5, the first functionally characterized three-finger protein (TFP) from the starfish Asterias rubens. Methods: Spatial structure determination was performed using 2D and 3D homo- and heteronuclear NMR. Intramolecular dynamics data (mobility in the pico-nanosecond and micro-millisecond ranges) were obtained by measuring 15N NMR relaxation parameters (R1, R2, and 15N-{1H}-NOE) with subsequent model-free analysis. Results and Discussion: The NMR data confirmed that Lystar5 adopts a classic "three-finger" fold stabilized by six disulfide bonds. The structure features two β-sheets: a small one in loop I and a larger one formed by four strands from all three loops, along with an α-helix in loop III. The molecular surface is mainly hydrophilic, containing two major clusters of negatively charged groups. Dynamics studies indicated that while the protein backbone is generally stable in the pico-nanosecond range, sections of loops I and II exhibit conformational exchange on the micro-millisecond timescale. Although the Lystar5 fold is similar to the human protein Lynx2, their distinct physicochemical properties and dynamics suggest different molecular targets and mechanisms of action. Conclusions: This work provides the first detailed structural and dynamic characterization of an echinoderm TFP. Understanding the relationship between the structure and dynamics of Lystar5 will facilitate the study of its interactions with human nicotinic acetylcholine receptors and integrins. These data open opportunities for the rational design of new biomedical drugs, specifically aimed at stimulating cell migration for wound healing.