Fluorescent detection of partially denatured conformation of beta-lactoglobuline

The native state of many proteins is known to be only marginally stable, correctly folded proteins are in the equilibrium with partially folded conformations and thus can convert to an aggregation-prone misfolded state [1]. Thus for globular proteins the partial unfolding is shown to be a first critical step in fibrillogenesis [2]. For fluorescent detection of globular proteins and their partially unfolded conformations "hydrophobic probes" anilinonaphthalene-sulfonate dyes such as ANS and bis-ANS are applied.

Series of tri and pentamethine cyanines are proposed as "external" fluorescent probes for globular proteins detection. The fluorescence sensitivity to partially folded conformations of amyloidogenic protein beta-lactoglobuline (BLG) was demonstrated by indolenine pentamethines with spatial hydrophobic substituent and benzothiazole squaraines (Fig.4). These dyes bind to BLG and increase their emission intensity upon temperature-induced unfolding of this protein. Thus, the dyes are able to monitor conformational changes of the protein.

Figure 4. Dyes sensitive to partially unfolded conformations of beta-lactoglobuline.

• C. M. Dobson Protein folding and misfolding. Nature 426, 884-890, (2003)
• V.Uversky, A. Finka Biochimica et Biophysica Acta 1698, 131– 153, (2004)
• Using of pentamethine dye 2904y for the detection of conformational changes of beta-lactoglobuline / V. B. Kovalska, M. Yu. Losytskyy, Yu. L. Slominskii, S. M. Yarmoluk // Lumcos 2011, Kharkov, November 2011 Book of Abstracts. P.76
• V.B.Kovalska, M. Yu. Losytskyy, P.F.Santos and S.M. Yarmoluk. .Fluorescent detection of partially denatured conformation of beta-lactoglobuline using squaraine dyes. 9-th International IUPAC Conference Polysolvat-9, Kyiv September 2012. Book of Abstracts. P.70