Influence of Microbial Transglutaminase on Physicochemical and Cross-Linking Characteristics of Individual Caseins

Abstract

The effects of microbial transglutaminase (MTGase) cross-linking on the physicochemical characteristics of individual caseins were investigated. MTGase was used to modify three major individual caseins, namely, κ-casein (κ-CN), αS-casein (αS-CN) and β-casein (β-CN). The SDS-PAGE analysis revealed that MTGase-induced cross-linking occurred during the reaction and that some components with high molecular weights (>130 kDa) were formed from the individual proteins κ-CN, αS-CN and β-CN. Scanning electron microscopy (SEM) and particle size analysis respectively demonstrated that the κ-CN, αS-CN and β-CN particle diameters and protein microstructures were larger and polymerized after MTGase cross-linking. The polymerized κ-CN (~749.9 nm) was smaller than that of β-CN (~7909.3 nm) and αS-CN (~7909.3 nm). The enzyme kinetics results showed KM values of 3.04 × 10−6, 2.37 × 10−4 and 8.90 × 10−3 M for κ-CN, αS-CN and β-CN, respectively, and, furthermore, kcat values of 5.17 × 10−4, 1.92 × 10−3 and 4.76 × 10−2 1/s, for κ-CN, αS-CN and β-CN, respectively. Our results revealed that the cross-linking of β-CN catalyzed by MTGase was faster than that of αS-CN or κ-CN. Overall, the polymers that formed in the individual caseins in the presence of MTGase presented a higher molecular weight and larger particles.

Impact of microbial transglutaminase and cooking time on functional properties of Mozzarella cheese analogues

“…Studies have suggested that these newly formed polymers are in direct correlation with reductions in as-caseins and bcaseins, confirming their reactivity with TGase and their affinity to form bonds with other caseins and serum proteins in the newly formed polymers (Romeih and Hamad 2010;Cadavid et al 2019;Abou-Soliman et al 2020;Chen et al 2020;Romeih et al 2021). However, this is in contrast with the findings of this study, where increased TGase concentration and cooking time resulted in casein (as, b and j) bands of higher intensity.…”

“…2020), although some studies have also found that α‐lactalbumin reduces with increases in TGase concentration as well (Chen et al . 2020).…”

“…The increased cooking time may have nullified the indigenous TGase inhibitors present in the milk serum or it may have modified the b-lactoglobulin through heat denaturation, causing them to unfold and expose the crosslinking sites to the TGase (Romeih and Walker 2017;Abou-Soliman et al 2020). Decreased b-lactoglobulin and increased a-lactalbumin whey protein banding with increased TGase concentration were reported by other literature (Cadavid et al 2020), although some studies have also found that a-lactalbumin reduces with increases in TGase concentration as well (Chen et al 2020).…”

“…TGase cross-links as-, band j-caseins, forming bonds between and within casein micelles creating a curd similar to that made by traditional methods (Aaltonen and Huumonen 2014). These crosslinking reactions can significantly change the protein molecules and lead to better texture, stability, water-holding capacity, emulsifying potential and gelation without any adverse effects on sensory or nutritional quality (Ercili-Cura and Huppertz 2015; Gharibzahedi et al 2018;Chen et al 2020).…”

Ultrasound-Assisted Transglutaminase Catalysis of the Cross-Linking and Microstructure of αs-Casein, β-Casein and κ-Casein

“…We noticed that the particles of β-CN with TGase and UT decreased from 15% to 0% yield for particles of approximately 400-6000 nm and increased from 0% to 13% for particles of approximately 6000-10,000 nm after 60 min of incubation (Figure 5C,D). Chen et al (2020) reported that it is the most sensitive to the response induced by TGase [32]. Furthermore, the particle size of κ-CN with TGase and UT decreased from a 4% to 2% yield for particles of approximately 100-6000 nm and increased from a 1% to 2% yield for particles of approximately 6000-10,000 nm, after 60 min of incubation (Figure 5E,F).…”