Structural studies of hemoglobin from two flightless birds, ostrich and turkey: insights into their differing oxygen-binding properties.

Crystal structures of hemoglobin (Hb) from two flightless birds, ostrich (Struthio camelus) and turkey (Meleagris gallopova), were determined. The ostrich Hb structure was solved to a resolution of 2.22 Å, whereas two forms of turkey Hb were solved to resolutions of 1.66 Š(turkey monoclinic structure; TMS) and 1.39 Š(turkey orthorhombic structure; TOS). Comparison of the amino-acid sequences of ostrich and turkey Hb with those from other avian species revealed no difference in the number of charged residues, but variations were observed in the numbers of hydrophobic and polar residues. Amino-acid-composition-based computation of various physical parameters, in particular their lower inverse transition temperatures and higher average hydrophobicities, indicated that the structures of ostrich and turkey Hb are likely to be highly ordered when compared with other avian Hbs. From the crystal structure analysis, the liganded state of ostrich Hb was confirmed by the presence of an oxygen molecule between the Fe atom and the proximal histidine residue in all four heme regions. In turkey Hb (both TMS and TOS), a water molecule was bound instead of an oxygen molecule in all four heme regions, thus confirming that they assumed the aqua-met form. Analysis of tertiary- and quaternary-structural features led to the conclusion that ostrich oxy Hb and turkey aqua-met Hb adopt the R-/RH-state conformation.

SARS-CoV-2 spike protein unlikely to bind to integrins via the Arg-Gly-Asp (RGD) motif of the Receptor Binding Domain: evidence from structural analysis and microscale accelerated molecular dynamics

The Receptor Binding Domain (RBD) of SARS-CoV-2 virus harbors a sequence of Arg-Gly-Asp tripeptide named RGD motif, which has also been identified in extracellular matrix proteins that bind integrins as well as other disintegrins and viruses. Accordingly, integrins have been proposed as host receptors for SARS-CoV-2. The hypothesis was supported by sequence and structural analysis. However, given that the microenvironment of the RGD motif imposes structural hindrance to the protein-protein association, the validity of this hypothesis is still uncertain. Here, we used normal mode analysis, accelerated molecular dynamics microscale simulation, and protein-protein docking to investigate the putative role of RGD motif of SARS-CoV-2 RBD for interacting with integrins. We found, by molecular dynamics, that neither RGD motif nore its microenvironment show any significant conformational shift in the RBD structure. Highly populated clusters were used to run a protein-protein docking against three RGD-binding integrin types, showing no capability of the RBD domain to interact with the RGD binding site. Moreover, the free energy landscape revealed that the RGD conformation within RBD could not acquire an optimal geometry to allow the interaction with integrins. Our results highlighted different structural features of the RGD motif that may prevent its involvement in the interaction with integrins. We, therefore, suggest, in the case where integrins are confirmed to be the direct host receptors for SARS-CoV-2, a possible involvement of other residues to stabilize the interaction.

Crystal structure of hemoglobin from mouse (Mus musculus) compared with those from other small animals and humans.

Mice (Mus musculus) are nocturnal small animals belonging to the rodent family that live in burrows, an environment in which significantly high CO2 levels prevail. It is expected that mouse hemoglobin (Hb) plays an important role in their adaptation to living in such a high-CO2 environment, while many other species cannot. In the present study, mouse Hb was purified and crystallized at a physiological pH of 7 in the orthorhombic space group P212121; the crystals diffracted to 2.8 Šresolution. The primary amino-acid sequence and crystal structure of mouse Hb were compared with those of mammalian Hbs in order to investigate the structure-function relationship of mouse Hb. Differences were observed from guinea pig Hb in terms of amino-acid sequence and from cat Hb in overall structure (in terms of r.m.s.d.). The difference in r.m.s.d. from cat Hb may be due to the existence of the molecule in a conformation other than the R-state. Analysis of tertiary- and quaternary-structural features, the α1ß2 interface region and the heme environment without any ligands in all four heme groups showed that mouse methemoglobin is in an intermediate state between the R-state and the T-state that is much closer to the R-state conformation.