The Conformation of Interfacially Adsorbed Ranaspumin-2 is an Arrested State on the Unfolding Pathway

TL;DR

This study combines experiments and simulations to reveal that Ranaspumin-2 adsorbs at interfaces through an unusual 'unhinging' process, maintaining secondary structure while adopting an arrested unfolded state, advancing understanding of protein-interface interactions.

Contribution

The paper introduces a modified Gō-model with explicit side-chains to accurately simulate Rsn-2's interfacial behavior and confirms the clam-shell unhinging mechanism experimentally.

Findings

Rsn-2 adsorbs via a two-step process involving a hydrophobic tail and unfolding.

The interfacial conformation is an arrested state along the unfolding pathway.

Simulations support the clam-shell unhinging model and show maintained secondary structure.

Abstract

Ranaspumin-2 (Rsn-2) is a surfactant protein found in the foam nests of the t\'{u}ngara frog. Previous experimental work has led to a proposed model of adsorption which involves an unusual clam shell-like `unhinging' of the protein at an interface. Interestingly, there is no concomitant denaturation of the secondary structural elements of Rsn-2 with the large scale transformation of its tertiary structure. In this work we use both experiment and simulation to better understand the driving forces underpinning this unusual process. We develop a modified G\={o}-model approach where we have included explicit representation of the side-chains in order to realistically model the interaction between the secondary structure elements of the protein and the interface. Doing so allows for the study of the underlying energy landscape which governs the mechanism of Rsn-2 interfacial adsorption. Experimentally, we study targeted mutants of Rsn-2, using the Langmuir trough, pendant drop tensiometry and circular dichroism, to demonstrate that the clam-shell model is correct. We find that Rsn-2 adsorption is in fact a two-step process: the hydrophobic N-terminal tail recruits the protein to the interface after which Rsn-2 undergoes an unfolding transition which maintains its secondary structure. Intriguingly, our simulations show that the conformation Rsn-2 adopts at an interface is an arrested state along the denaturation pathway. More generally, our computational model should prove a useful, and computationally efficient, tool in studying the dynamics and energetics of protein-interface interactions.