Towards resolving the complex paramagnetic nuclear magnetic resonance (NMR) spectrum of small laccase: assignments of resonances to residue-specific nuclei
| Autor: | H. J. M. de Groot, K.B. Sai Sankar Gupta, Rubin Dasgupta, Marcellus Ubbink |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
0301 basic medicine
QC501-766 biology Ligand chemistry.chemical_element Active site Nuclear magnetic resonance spectroscopy 010402 general chemistry 01 natural sciences Copper 0104 chemical sciences Electricity and magnetism NMR spectra database 03 medical and health sciences chemistry.chemical_compound Paramagnetism Crystallography 030104 developmental biology chemistry biology.protein Imidazole Histidine |
| Zdroj: | Magnetic Resonance Magnetic Resonance, Vol 2, Pp 15-23 (2021) |
| ISSN: | 2699-0016 |
| Popis: | Laccases efficiently reduce dioxygen to water in an active site containing a tri-nuclear copper centre (TNC). The dynamics of the protein matrix is a determining factor in the efficiency in catalysis. To probe mobility, nuclear magnetic resonance (NMR) spectroscopy is highly suitable. However, several factors complicate the assignment of resonances to active site nuclei in laccases. The paramagnetic nature causes large shifts and line broadening. Furthermore, the presence of slow chemical exchange processes of the imidazole rings of copper ligand results in peak doubling. A third complicating factor is that the enzyme occurs in two states, the native intermediate (NI) and resting oxidized (RO) states, with different paramagnetic properties. The present study aims at resolving the complex paramagnetic NMR spectra of the TNC of Streptomyces coelicolor small laccase (SLAC). With a combination of paramagnetically tailored NMR experiments, all eight His Nδ1 and Hδ1 resonances for the NI state are identified, as well as His Hβ protons for the RO state. With the help of second-shell mutagenesis, selective resonances are tentatively assigned to the histidine ligands of the copper in the type-2 site. This study demonstrates the utility of the approaches used for the sequence-specific assignment of the paramagnetic NMR spectra of ligands in the TNC that ultimately may lead to a description of the underlying motion. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|