Profiling of dynamics in protein–lipid–water systems: a time-resolved fluorescence study of a model membrane protein with the label BADAN at specific membrane depths

Abstract

Profiles of lipid-water bilayer dynamics were determined from picosecond time-resolved fluorescence spectra of membrane-embedded BADAN-labeled M13 coat protein. For this purpose, the protein was labeled at seven key positions. This places the label at well-defined locations from the water phase to the center of the hydrophobic acyl chain region of a phospholipid model membrane, providing us with a nanoscale ruler to map membranes. Analysis of the time-resolved fluorescence spectroscopic data provides the characteristic time constant for the twisting motion of the BADAN label, which is sensitive to the local flexibility of the protein–lipid environment. In addition, we obtain information about the mobility of water molecules at the membrane–water interface. The results provide an unprecedented nanoscale profiling of the dynamics and distribution of water in membrane systems. This information gives clear evidence that the actual barrier of membranes for ions and aqueous solvents is located at the region of carbonyl groups of the acyl chains.

Spectral properties of BADAN in solutions with different polarities

“…Fluorescent solvatochromic dye BADAN (6-bromo-acetyl-2-dimethylamino-naphtalene) ( Figure 1) is used for studying of the phase transitions in biological membranes, protein conformational changes induced by different factors, proteins interactions with their partners and ligands [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15]. Furthermore, fluorescence of this dye is used as a recorded signal in biosensor systems [16][17][18][19][20][21].…”

“…The dependence of BADAN fluorescent characteristics on its environment properties is often explained by use the data about its well-studied structural analogues PRODAN and LAURDAN [4,8,9]. The fluorescence of these dyes has dual character and is described in terms of locally excited state and charge-transfer state in polar solvents [24][25][26][27][28].…”

Increased Structural Flexibility at the Active Site of a Fluorophore-conjugated β-Lactamase Distinctively Impacts Its Binding toward Diverse Cephalosporin Antibiotics

“…For our design we decided to use the naphthalene-type fluorophore molecules because of their exceptional solvachromatic properties that can reflect the polarity of the local environmental at the molecule level, as well as their relative small size comparable to the amino acid, tryptophan, to allow them to be positioned close to the protein structure (7)(8)(9)(10)(11)(12)(13). This fluorophore has been applied to probe the structural dynamics of membrane lipids (11) and ion channels (12), and to detect the binding of peptide ligands to SH2 domains (9) and Class II MHC proteins (13).…”

“…This fluorophore has been applied to probe the structural dynamics of membrane lipids (11) and ion channels (12), and to detect the binding of peptide ligands to SH2 domains (9) and Class II MHC proteins (13). We examined several fluorophores and report here one such engineered ␤-lactamase, PenP-E166Cb, which is based on the Class A ␤-lactamase PenP from Bacillus licheniformis.…”

Reverse micelles as a tool for probing solvent modulation of protein dynamics: Reverse micelle encapsulated hemoglobin

“…The spectra presented are normalized with respect to intensity. The actual emission intensity increases dramatically as the peak undergoes shifting to the blue [55]. Fig.…”