Cross-linking with microbial transglutaminase: Isopeptide bonds and polymer size as drivers for acid casein gel stiffness

Addition of crude tiger nut protein extract affects stiffness of enzymatically cross‐linked dairy proteins

“…Increasing gel stiffness upon enzymatic cross‐linking of sodium caseinate was attributed to the formation of covalent isopeptide bonds. However, larger casein polymers may lack in the molecular flexibility that is necessary to form stiffer gels (Raak et al ., , ). Addition of crude tiger nut proteins containing 2.8 g kg −1 minerals to sodium caseinate before enzymatic cross‐linking (CTe) may have affected the ionic characteristics of the milieu and favoured protein association during cross‐linking, contributing to a more pronounced decrease in gel stiffness because of the formation of larger polymers.…”

“…On the other hand, WTe showed higher gel stiffness than WeT although there was no significant difference in their DPs (WTe, 33.12 AE 0.10%; WeT, 35.09 AE 0.74%). Polymer size and isopeptide content, that is the amount of intermolecular and intramolecular crosslinks, are known to affect the molecular flexibility and compactness of polymers, thereby influencing gel stiffness of enzymatically cross-linked dairy proteins (Raak et al, 2017).…”

Size Separation Techniques for the Characterisation of Cross-Linked Casein: A Review of Methods and Their Applications

“…Peptide bonds between terminal functional groups, referred to as isopeptide bonds, are not susceptible to most proteolytic enzymes and remain therefore intact after total enzymatic protein hydrolysis; subsequent amino acid analysis by chromatographic techniques enables the determination of the N-ε-(γ-glutamyl)-lysine isopeptide [29]. Although the isopeptide content was shown to be an important driver for the stiffness of acid-induced casein gels, polymer size still has to be taken into account [30][31][32]. Besides TGase, oxidoreductases were applied, which have a completely different cross-linking mechanism: firstly, the enzymes oxidise specific functional groups that react further in non-enzymatic reactions.…”

“…For instance, the highest number of casein dimers in TGase-catalysed polymerisation was observed in moderately cross-linked samples, i.e., at high levels of residual monomers (>50%), indicating that dimers as well as smaller oligomers are good substrates for further cross-linking by TGase [32,78,146]. Moreover, the polymerisation degree (sometimes also referred to as oligomerisation degree) was calculated in several studies from size exclusion chromatograms by relating peak areas of cross-linked caseins to the entire sample area [30,32,44,47,48,53,78,[134][135][136][146][147][148]. Because reference casein samples often reveal a low level of polymerised casein (see Figures 9 and 10), the polymerisation degree after cross-linking was also expressed as the difference between cross-linked samples and reference [137][138][139]144,145].…”

The Use of Trisodium Citrate to Improve the Textural Properties of Acid-Induced, Transglutaminase-Treated Micellar Casein Gels

“…The primary building blocks of acid-induced milk gel network structures are proteins. Microbial transglutaminase (mTGase) is an enzyme widely used to form covalent links between individual protein molecules [ 3 , 4 , 5 , 6 ]. The textural properties of acid-induced gels prepared from mTGase-treated milk proteins can be significantly enhanced [ 2 , 7 ].…”