Cytochrome b561 proteins (CYB561s) are integral membrane proteins with six trans-membrane domains, two heme-b redox centers, one on each side of the host membrane. The major characteristics of these proteins are their ascorbate reducibility and trans-membrane electron transferring capability. More than one CYB561 can be found in a wide range of animal and plant phyla and they are localized in membranes different from the membranes participating in bioenergization. Two homologous proteins, both in humans and rodents, are thought to participate—via yet unidentified way—in cancer pathology. The recombinant forms of the human tumor suppressor 101F6 protein (Hs_CYB561D2) and its mouse ortholog (Mm_CYB561D2) have already been studied in some detail. However, nothing has yet been published about the physical-chemical properties of their homologues (Hs_CYB561D1 in humans and Mm_CYB561D1 in mice). In this paper we present optical, redox and structural properties of the recombinant Mm_CYB561D1, obtained based on various spectroscopic methods and homology modeling. The results are discussed in comparison to similar properties of the other members of the CYB561 protein family.

The accumulation of proteins in filter membranes limits the efficiency of filtering technologies for cleaning wastewater. Efforts are ongoing to coat commercial filters with different materials (such as titanium dioxide, TiO₂) to reduce the fouling of the membrane. Beyond monitoring the desired effect of the retention of biomolecules, it is necessary to understand what the biophysical changes are in water-soluble proteins caused by their interaction with the new coated filter membranes, an aspect that has received little attention so far. Using spin-label electron paramagnetic resonance (EPR), aided with native fluorescence spectroscopy and dynamic light scattering (DLS), here, we report the changes in the structure and dynamics of bovine serum albumin (BSA) exposed to TiO₂ (P25) nanoparticles or passing through commercial polyvinylidene fluoride (PVDF) membranes coated with the same nanoparticles. We have found that the filtering process and prolonged exposure to TiO₂ nanoparticles had significant effects on different regions of BSA, and denaturation of the protein was not observed, neither with the TiO₂ nanoparticles nor when passing through the TiO₂-coated filter membranes.

Accumulation of proteins in filter membranes limits the efficiently of filtering technologies for cleaning wastewater. Efforts are ongoing to coat commercial filters with different materials (such as titanium-dioxide, TiO2) in order to reduce the fouling of the membrane. Beyond monitoring the desired effect of retention of biomolecules, it is demanding to understand what are the biophysical changes in water-soluble proteins caused by interacting with the new coated filter membranes, an aspect that has received little attention so far. Using spin label electron paramagnetic resonance (EPR), aided with native fluorescent spectroscopy and dynamic light scattering (DLS), here we report the changes in the structure and dynamics of bovine serum albumin (BSA) exposed to TiO2 (P25) nanoparticles or passing through commercial polyvinylidene fluoride (PVDF) membranes coated with the same nanoparticles. We have found that the filtering process and prolongued exposure to TiO2 nanoparticles had significant effects on different regions of BSA, and denaturation of the protein was not observed, neither with TiO2 nanoparticles nor by pressing through TiO2-coated filter membranes.

The lipid composition of thylakoid membranes (TMs), the site of light reactions in oxygenic photosynthetic organisms, is dominated by the non-bilayer lipid species monogalactosyldiacylglycerol. It has been documented that plant TMs display strong lipid polymorphisms (Garab et al. 2022 Prog Lipid Res 86:101163). Recently, we have also shown that BBY membrane sheets, large, laterally fused photosystem-II (PSII) enriched membrane pairs, beside the lamellar phase, contain an intense isotropic phase, the lipid molecules of which mediate the fusion of membranes (Böde et al. 2024 Photosynth Res 161:127-140). To demonstrate the composite nature of BBY, we employed confocal laser scanning microscopy, using the lipid-label BODIPY-phosphatydilcholine and chlorophyll-a fluorescence emissions. To characterize the physico-chemical microenvironments of lipid molecules, we stained BBY membranes with the hydrophobic fluorescent dye DPH (1,6-diphenyl-1,3,5-hexatriene). DPH emission spectra from face- and edge-aligned BBY membranes indicated the existence of at least two distinct microenvironments. Fluorescence lifetime analyses revealed three components; the fastest one was sensitive to the enzymatic treatment with wheat germ lipase (WGL), which had earlier been shown to selectively eliminate the isotropic lipid phase of BBY and to disassemble the large sheets into its constituent membrane pairs of grana. Although the anisotropic fluorescence decay kinetics discerned no lifetime heterogeneity of the untreated DPH-stained BBY, WGL treatment led to the appearance of a second lifetime component. These data provide independent experimental evidence on the lipid polymorphism of BBY membranes and reveal that the bilayer lipids and the non-bilayer lipid arrays mediating the fusion of TMs possess distinct physico-chemical environments.