Site-Selective Dynamics of Azidolysozyme

TL;DR

This study investigates the local dynamics of azido-modified alanine residues in lysozyme using spectroscopy, revealing their sensitivity to local environment and hydration effects, with implications for probing protein dynamics.

Contribution

It provides detailed characterization of site-specific dynamics of azido-modified alanine residues in lysozyme, highlighting their sensitivity and minimal structural perturbation.

Findings

AlaN₃ is a positionally sensitive probe for local dynamics.

Frequency span of AlaN₃ response is ~15 cm⁻¹, consistent with other amino acid replacements.

Static component of dynamics correlates with hydration level around the probe.

Abstract

The spectroscopic response of and structural dynamics around all azido-modified alanine residues (AlaN$_3$) in Lysozyme is characterized. It is found that AlaN$_3$ is a positionally sensitive probe for the local dynamics, covering a frequency range of $\sim 15$ cm$^{-1}$ for the center frequency of the line shape. This is consistent with findings from selective replacements of amino acids in PDZ2 which reported a frequency span of $\sim 10$ cm$^{-1}$ for replacements of Val, Ala, or Glu by azidohomoalanine (AHA). For the frequency fluctuation correlation functions (FFCFs) the long-time decay constants $\tau_2$ range from $\sim 1$ to $\sim 10$ ps which compares with experimentally measured correlation times of 3 ps. Attaching azide to alanine residues can yield dynamics that decays to zero on the few ps time scale (i.e. static component $\Delta_0 \sim 0$ ps$^{-1}$) or to a remaining, static contribution of $\sim 0.5$ ps$^{-1}$ (corresponding to 2.5 cm$^{-1}$), depending on the local environment on the 10 ps time scale. The magnitude of the static component correlates qualitatively with the degree of hydration of the spectroscopic probe. Although attaching azide to alanine residues is found to be structurally minimally invasive with respect to the overall protein structure, analysis of the local hydrophobicity indicates that the hydration around the modification site differs for modified and unmodified alanine residues, respectively.