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bioRxiv · 10.1101/2020.06.09.142257

Real-Time Microfluidics-Magnetic Tweezers connects conformational stiffness with energy landscape by a single experiment

chakraborty, s.·Chaudhuri, D.·Haldar, S.·Singh, V.·Banerjee, S.
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Abstract

Proteins are versatile biopolymers whose functions are determined by their structures. Understanding the structural dynamicity, with respect to energy landscape, is essential to describe their biological functions. The ability to study the dynamical properties of a single protein molecule is thus crucial, but ensuring that multiple physical properties can be simultaneously extracted within a single experiment on the exact same protein molecule in real-time has hitherto been infeasible. Here, we present magnetic tweezers technology that surmounts this limitation, providing real-time dynamic information about changes in several molecular properties ({Delta}G0, conformation, and mean first passage time of unfolding and refolding) from a single experiment, by remeasuring the very same protein molecule in varying chemical condition. We illustrate the versatility of the method by studying electrolyte-dependent conformational flexibility and the energy landscape of substrate protein L under force. Changing salt concentrations reshapes the energy landscape by two specific ways: it speeds-up refolding kinetics while slowing down unfolding kinetics. From the same trajectory, we calculate the stiffness of the protein polymer, a quantity that varies with salt concentration. The data is described within the framework of a modified electrolyte FJC model that we propose and study here. The observed correlation between{Delta} G0, kinetics and polymer elasticity connect protein chain physics and the energy landscape, while the experimental methodology we describe of getting energy landscape from a single experiment could have wide-ranging applications.

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chakraborty, s., Chaudhuri, D., Haldar, S., Singh, V., Banerjee, S.. 2020-06-10. Real-Time Microfluidics-Magnetic Tweezers connects conformational stiffness with energy landscape by a single experiment. https://doi.org/10.1101/2020.06.09.142257

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