Addressing Mechanism of Fibrillization/Aggregation and Its Prevention in Presence of Osmolytes: Spectroscopic and Calorimetric Approach

Abstract

Understanding the mechanism of protein fibrillization/aggregation and its prevention is the basis of development of therapeutic strategies for amyloidosis. An attempt has been made to understand the nature of interactions of osmolytes L-proline, 4-hydroxy-L-proline, sarcosine and trimethylamine N-oxide with the different stages of fibrillization of hen egg-white lysozyme by using a combination of isothermal titration calorimetry, differential scanning calorimetry, fluorescence spectroscopy, and transmission electron microscopy. Based on thioflavin T fluorescence emission intensities and microscopic images, the nucleation, elongation, and saturation phases of fibrillization have been identified. Isothermal titration calorimetry and differential scanning calorimetry have enabled a quantitative analysis of the nature of interactions of these osmolytes with various conformational states of lysozyme at different stages of fibrillization/aggregation. It is concluded that interaction of the osmolytes with lysozyme fibrils at both the nucleation and elongation stages are important steps in the prevention of fibrillization/aggregation. Identification of the nature of interactions is a key step towards the discovery and synthesis of target oriented potential inhibitors of these associations. This study is a first report in which calorimetry has been used to address interaction of potential inihibitiors with the protein at different stages of fibrillization.

ITC and SPR Analysis Using Dynamic Approach

“…Yet again the advantage of dynamic approach is that the instrument response can be seamlessly integrated within the kinetic framework of the binding mechanism thereby simplifying the data analysis. A representative of different kinds of binding mechanisms such as, single set of equivalent sites, two sets of equivalents sites with sequential binding mode, two sets of equivalents sites with parallel binding mode, and aggregation have been considered here [15,16,17,18,19,20,21,22,23,24,25,26]. The simulation of ITC profiles for all these mechanisms were realised and found to be consistent with that of the previous reports [8].…”

“…Aggregation mechanisms are usually complex and difficult to model, due to the various binding modes available for the intra and inter molecular complex formation [24,26,35]. However, any complex aggregation mechanism can be modelled as a series of sequential binding mechanism, with several steps in between the monomer and the final aggregate polymer.…”

“…In the all the numerical simulations mentioned in this work we used the 'difference' form of the above equations Eqn( 21,22) rather than its differential form, as shown below Eqns (23,24),…”

Conditions Governing Food Protein Amyloid Fibril Formation—Part I: Egg and Cereal Proteins

“…Numerous researchers have investigated the inhibition of LYS amyloid formation. This has resulted in a list of compounds that either partially or completely inhibit fibril formation at acidic pH: food constituents such as short‐chain phospholipids (pH 2.0 and 55 °C) (Wang, Hung, Wen, Lin, & Chen, ), curcumin (Liu et al., ; Wang, Liu, & Lee, ) or its water‐soluble derivatives (Wang et al., ), myricetin (He et al., ), the osmolytes proline, hydroxyproline, sarcosine and trimethylamine N‐oxide (Choudhary & Kishore, ), cysteine (Takai et al., ; Wang, Liu, Wu, & Lai, ), glutathione (Wang, Chou, Liu, & Wu, ), carnosine (Wu et al., ), trehalose, magnesium chloride (Chatterjee, Kolli, & Sarkar, ), safranal, crocin (Joloudar et al., ), zinc ions (Ma, Zhang, Wang, & Zhu, ), aroma components (for example, phenyl ethyl alcohol, N,N,N,N’‐tetramethylethylenediamine, or cinnamaldehyde) (Seraj et al., ), rosmarinic acid, resveratrol (Shariatizi, Meratan, Ghasemi, & Nemat‐Gorgani, ), chemicals such as p‐benzoquinone (Lieu et al., ; Wang, Chen, & Hung, ), 4‐aminophenol and 2‐amino‐4‐chlorophenol (Vieira, Figueroa‐Villar, Meirelles, Ferreira, & De Felice, ), tris(2‐carboxyethyl)phosphine (Wang, Liu, & Lu, ), SDS concentrations of at least 0.25 mM (Hung et al., ), nonionic detergents like triton X‐100 and n‐dodecyl‐β‐D‐maltoside (Siposova, Kozar, & Musatov, ), the ionic liquid tetramethyl guanidinium acetate (Kalhor, Kamizi, Akbari, & Heydari, ), clotrimazole (Sarkar, Kumar, & Dubey, ), sodium tetrathionate (Sarkar, Kumar, & Dubey, ), β‐mercaptoethanol (Sarkar et al., ), glycol‐acridines (Vuong et al., ), 2‐acetyl amin‐3‐[4‐(2‐amintgo‐5‐sulfo‐phenyl]‐propionic acid (Maity et al., ), indole, indole 3‐acetic acid, indole 3‐carbinol, indole 3‐propionic acid and tryptophol (Morshedi, Rezaei‐Ghaleh, Ebrahim‐Habibi, Ahmadian, & Nemat‐Gorgani, ), and other components such as melatonin (Wang, Chen, et al., ), glutathione‐covered gold nanoparticles (Antosova et al., ), manganese–salen derivatives (Bahramikia & Yazdanparast, ; Bahramikia, Yazdanparast, & Gheysarzadeh, ), the crowding agents Ficoll 70 and dextran 70 (Ma et al., ), bovine serum albumin and its combination with Ficoll 70 (Zhou, Zhou, Hu, Chen, & Liang, ), and type I collagen (Dubey & Mar, ).…”

Biological Wonders of Osmolytes: The Need to Know More

“…Protein fibrillation is responsible for several amyloidogenic disorders including diseases such as Alzheimer's, Parkinson's, mad cow, diabetes type II, cystic fibroisis, and dialysis related amyloidosis [58,59]. Though there have been several studies describing the effects of osmolytes on fibrillation/aggregation of proteins [60][61][62][63][64][65][66], quantitative understanding in terms of energetics of interaction has only recently been addressed Page 3 of 5 [67,68]. Trehalose has been currently used for the treatment of Huntington's disease in transgenic animal mice [69].…”