A multifrequency HYSCORE study of weakly coupled nuclei in frozen solutions of high-spin aquometmyoglobin.

The research represents advancements in use of multifrequency HYSCORE (Hyperfine Sublevel Correlation Spectroscopy) to dissect the way electron spin interacts with remote nuclei in ferric heme proteins, using Horse heart aquo-metmyoglobin as a model.

Introduction

• The researchers demonstrate the efficiency of multifrequency HYSCORE (Hyperfine Sublevel Correlation Spectroscopy), a technique used to investigate the hyperfine couplings between electron spins and nucleic spins, in the study of ferric heme proteins.
• In this study, Horse heart aquo-metmyoglobin was chosen as the model system for the application of this advanced method.

Methods and Findings

• Using these techniques, experimental evidence was collected and assigned to various components such as the protons of the proximal histidine ligand, the mesoprotons of the heme ligand, and two different protons of the distal water ligand.
• The researchers registered HYSCORE signals from the remote N(delta) of both proximal and distal histidines, proving the sensitivity and efficiency of the HYSCORE techniques.
• Remarkably, the study was even able to detect correlation peaks from the less-abundant (13)C nuclei of the heme ligand.

Implications

• This investigation led to completely new findings that allow for a detailed understanding of the hyperfine couplings at the individual level and the computation of the spin density in the pi and sigma orbitals.
• The texture of information acquired from this study rewards previous research data obtained from both the electron paramagnetic resonance and liquid-state NMR techniques, further enhancing the understanding of ferric heme proteins.