Enzymatic Cross-Linking of β-Lactoglobulin:  Conformational Properties Using FTIR Spectroscopy

Abstract

In this study, we use FTIR spectroscopy to probe the conformational changes of beta-lactoglobulin (beta-LG)-the main constituent of whey proteins-as subjected to enzymatic cross-linking by transglutaminase. We investigate both the amide I region (1600-1700 cm(-1)) and the C-H stretching region (2800-3100 cm(-1)). In the amide I region, spectra of denatured conformations of beta-LG, known to be necessary for cross-linking, differ according to the denaturation procedure, i.e., chemical or thermal treatment. Denaturation by chemical denaturants, dithiothreitol (DTT) or beta-mercaptoethanol, show no effect on the alpha-helix, while shifting the monomer dimer equilibrium toward higher monomer concentration. On the other hand, denaturing by thermal treatment dissociates the beta-sheets in the native structure, leading to new intermolecular beta-sheets being formed. Preheated then enzyme cross-linked beta-LG molecules show very similar spectra in the amide I region to the molecules with no cross-linking, indicating minimal effects of the cross-links on the carbonyl stretching mode. However, chemically denatured (using beta-mercaptoethanol) then enzyme cross-linked beta-LG molecules show noticeable diminution in the alpha-helix band and formation of strong hydrogen-bonded intermolecular beta-sheets. In the C-H stretching region, preheated then enzyme cross-linked beta-LG molecules exhibit a different degree of exposure of aliphatic amino acids due to the enzyme action. The same behavior is observed for DTT-treated then enzyme cross-linked beta-LG molecules. Generally, the changes in the C-H stretching region clearly indicate that hydrophobic interactions are altered upon enzymatic cross-linking.

Physicochemical properties and microstructure of soy protein hydrogels co-induced by Maillard type cross-linking and salts

“…Scale bar: 100 nm. Kadla, & Khan, 2006). These methods of gelation do not lead to modification of protein secondary structure since cross-linking or addition of salt does not appear to influence hydrogen bonding.…”

Newtonian viscosity behavior of dilute solutions of polymerized whey proteins. Would viscosity measurements reveal more detailed molecular properties?

“…The presence of molecular structuring was confirmed above by the calculated thermodynamic parameters. Structuring at moderate to higher concentrations of whey protein polymers has been studied in previous work (Eissa, Puhl, Kadla, & Khan, 2006;Elofsson et al, 1996;Jost, 1995;Mulvihill & Donovan, 1987;Bolder, Vasbinder, Sagis, & van der Linden, 2007;Vardhanabhuti & Foegeding, 1999). However, at appreciably lower concentrations, similar to these concentrations used in DSV experiments, structuring has not been thoroughly discussed.…”

FTIR Spectroscopic Study of Heat-Induced β-Lactoglobulin Solution under Flow Field

“…In this amide I region (1700-1600 cm -1 ), the band appears at 1660-1640 cm -1 for α-helix and random coils and also band at 1640-1620 cm -1 appears for β-sheet. These bands in the FTIR spectra have also explained the secondary structure of the β-lactoglobulin [33][34][35][36]. Fig.…”