Neuroglobin (Ngb) is a new member of hexa-coordinat vertebrate Heme-globin with several proposed functions such as ROS/RNS scavenging, and inhibition of hypoxia-induced apoptosis. Nonetheless, the molecular mechanism for most of its functions is still not clear. Recent studies have shown a link between the structure and function of the protein, for example, the CD loop in Ngb is a potential binding site for the G<sub>α</sub> subunit and the distal histidine affects the thermodynamic profile for gaseous ligands association. In order to characterize the structure-function relationship in Ngb, we have introduced two mutants, the E53Q in the CD loop region and H64Q in the distal pocket. UV-vis and fluorescence spectroscopy together with time of flight-mass spectrometry have been used to determine how the mutations affect the Ngb stability. Our data indicate a distinct stability profile of the two mutants in both liquid phase and gas phase. In the gas phase, Ngb WT is more stable in neutral pH whereas stability of Ngb mutants was pH independent. However, both mutants, especially E53Q, are significantly less stable in the gas phase than the wild type. In the liquid phase, distal histidine replacement leads to a decrease in overall protein stability whereas the stability of E53Q mutant is comparable to that of WT. Nevertheless, the replacement of glutamine residue increases the protein solubility during pH induced unfolding studies. Overall, our data provide information on the impact of the CD loop and the distal histidine on protein stability in both gas phase and liquid phase.<br/>

Mutation of residues in CD loop and distal pocket impact protein stability of human neuroglobin

2019 Spring National Meeting

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