Microbial Transglutaminase Treatment Research Articles

Effect of Microbial Transglutaminase Treatment on the Techno-Functional Properties of Mung Bean Protein Isolate.

The purpose of this study was to investigate the improvement in techno-functional properties of mung bean protein isolate (MBPI) treated with microbial transglutaminase (MTG), including water- and oil-holding capacity, gelling properties, and emulsifying capacity. MBPI dispersions were incubated with MTG (5 U/g of protein substrate) at 45 °C with constant stirring for 4 h (MTM4) or 8 h (MTM8). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that MTG treatment for different durations increased the amount of high-molecular-weight proteins in MBPI, and most of the cross-linking by MTG was terminated at 8 h. Improved water-holding capacity, gelling properties, emulsifying capacity, and stability were observed after MTG treatment, and decreased protein solubility and surface hydrophobicity were observed. Furthermore, the texture of the heat-induced gels made from MTG-treated MBPI was evaluated using a texture analyzer. MTG treatment increased the hardness, gumminess, chewiness, and adhesiveness of the heat-induced gels. Field-emission scanning electron microscopy demonstrated the enhanced hardness of the gels. This research reveals that MTG-catalyzed cross-linking may adjust the techno-functional properties of MBPI, allowing it to be used as a soy protein alternative in food products, such as plant-based and processed meats.

Hofmeister anion effects synergize with microbial transglutaminase to enhance the techno-functional properties of pea protein

Pea protein are emerging as the most potential alternative for meat products, but its application was hindered by their weaker gelling properties. Here, the feasibility of combining the Hofmeister anion (CO32–, Citrate3-, and SO42-) effect with microbial transglutaminase (MTG) cross-linking strategy to improve the techno-functional properties of pea protein was studied. Hofmeister anions or/and MTG treatment of pea protein caused a clear shift in far-UV CD spectra towards β-turn and random coil structures. Furthermore, Hofmeister anion and MTG-induced crosslinking caused a reduction of surface hydrophobicity in contrast with anions-treated. Compared to CO32– and SO42-, Citrate3- treatment can better improve the efficiency of MTG-crosslinking, as demonstrated by a reduction in free amino group contents and an increase in mean diameter size. Using MTG in combination with Hofmeister anions showed significantly improved foam property and gel hardness as well as decrease gelation temperature of pea protein, specifically Citrate3- treatment. Thus, this research provides a novel and effective method to improve the effect of MTG-cross-linked pea protein, which will play an essential role in future food production.

Effect of storage time on sensory and instrumental properties of skim‐milk yoghurt obtained with microbial transglutaminase

Not many studies have examined changes in microbial transglutaminase (MTG) yoghurts during storage. In this work, yoghurts manufactured with skimmed milk and treated with MTG were evaluated in refrigerated storage for 35 days. Instrumental measurements showed that firmness, consistency and pH values in MTG yoghurt experienced a significant increase with the storage time. However, sensory measurements showed that MTG yoghurts were not affected significantly by storage time. Acid taste scores only increased significantly with storage time in the control samples. Microbial transglutaminase treatment showed a positive effect on textural properties and allowed syneresis to be prevented completely during storage.

Effects of high hydrostatic pressure and microbial transglutaminase treatment on structure and gelation properties of sweet potato protein

The effects of high hydrostatic pressure (HHP) combined with microbial transglutaminase (MTGase) on the structure and gelation properties of sweet potato protein (SPP) were investigated. MTGase induced the formation of high molecular mass aggregates of SPP by the inter-molecular cross-linking. The average particle size of SPP increased by HHP at 250–550 MPa, being increased from 249.5 nm (0.1 MPa) to 270.2 nm (550 MPa). While it was decreased by the subsequent MTGase catalysis that showed the lowest value of 200.3 nm at 550 MPa, indicating the formation of intra-molecular linkages. The amount of sulfhydryl groups and thermal denaturation temperature were increased by HHP and MTGase, with the highest value of 3.5 μmol/g protein and 80.4 °C in MTGase catalyzed SPP with 400 MPa pretreatment. Textural properties of the gels made from HHP treated SPP or those with further MTGase catalysis were both enhanced due to the covalent linkage of disulfide bonds (-S-S-) and ε-(γ-glutaminyl) lysine isopeptide bonds. In addition, low-field NMR results showed an increase in corresponding area fractions of A2b and A21, being attributed to more bound or immobilized water fractions were trapped in the SPP gel matrix.

Expression of recombinant transglutaminase gene in Pichia pastoris and its uses in restructured meat products

Transglutaminase is an effective enzyme that catalyzes the crosslinking of various meats, improves meat product quality, and is widely used in the meat industry. In this study, microbial transglutaminase (MTG) was expressed successfully in Pichia pastoris strain GS115, and the enzyme activity was approximately 0.70 U/ml. The recombinant MTG expressed in P. pastoris was used in the investigation of restructured pork and crosslinking of soy protein isolate (SPI) and chicken myofibrillar protein (MP). Results showed that the hardness, chewiness, and F1 of the restructured pork increased, and the adhesiveness decreased after MTG treatment, However, high temperature had greater effect on the texture of restructured pork after MTG treatment than that of the control. MTG can crosslink SPI component acidic subunits, subunits of β-conglycinin and MP component myosin heavy chain, and actin. MTG, as a food additive, can be successfully heterologously expressed, and the recombinant MTG has potential application in restructured meat products.

Effects of microbial transglutaminase treatment on physiochemical properties and emulsifying functionality of faba bean protein isolate

The potential use of microbial transglutaminase (MTG)-treated faba bean protein isolate (FBPI) as emulsifiers to maintain physical and oxidative stability of oil-in-water (O/W) emulsion was investigated. MTG-treated FBPIs (MTG-FBPIs) were prepared by incubating with MTG for 60, 120 or 240 min. O/W emulsions were stabilized by 3% (w/v) of MTG-FBPIs or control-FBPI (treated with inactive MTG) and stored at 37 °C for 7 days. MTG treatments induced cross-linking in FBPI, raised the protein net surface charges by 5%–8%, and increased the emulsion particle size by 19%–135%. MTG treatment for 120 and 240 min but not 60 min induced excessive surface hydrophobicity, resulting in decreased emulsifying activity and physical stability of emulsion. By day 7, all MTG-treated FBPIs showed similar inhibiting effects against lipid oxidation in emulsion, indicated by less conjugated dienes and hexanal production. MTG-FBPIs moderately promoted protein oxidation (120 min > 240 min ≈60 min). Thus, prolonged MTG treatment should be avoided to prevent accelerated protein oxidation and droplets coalescence. MTG treatment for 60 min makes FBPI a potential emulsifier to maintain physical stability while improving lipid oxidative stability in emulsion, potentially attributed to thicker interfacial layer, larger droplet size, and protective effect of protein.

Microbial transglutaminase enhances antioxidant activity of yogurt through altering pattern of water‐soluble peptides and increasing release of amino acids

SummaryThis study compared the antioxidant capacity of yogurt with and without microbial transglutaminase treatment after milk fermentation, and investigated the correlation between antioxidant property and the water‐soluble peptides and amino acids composition. Results showed that small molecular peptide fraction exhibited stronger antioxidant activity than large peptides. Microbial transglutaminase yogurt isolated fraction had stronger antioxidant capacity than that from the control. Microbial transglutaminase altered the peptides composition, and resulted in higher amount of small molecular peptides (<1.5 kDa) mainly consisting of κ‐casein‐derived peptides in yogurt. The dominant peptides sequences (IAKYI, IAKYIPI, IAKYIPIQY, PYYAK and NQFLPYPYYAK) in the treated yogurt were different from those in the control (PYPQ, VLPVPQKAVPYPQ, PVPQKAVPYPQ and VLSRYPSYGLN). Microbial transglutaminase improved the composition and concentration of amino acids in yogurt.

Microbial Transglutaminase Used in Bread Preparation at Standard Bakery Concentrations Does Not Increase Immunodetectable Amounts of Deamidated Gliadin

The effect of standard bakery concentrations of microbial transglutaminase (MTG) in wheat bread preparation on the immunoreactivity of sera of celiac disease (CD) patients was investigated. Immunoblotting using monoclonal antibodies specific to unmodified and/or deamidated gliadin showed no differences between control bread and MTG bread. Deamidation of gliadin could not be detected at standard MTG concentrations. Sera of CD patients were characterized using anti-gliadin and anti-deamidated gliadin peptide (DGP) enzyme-linked immunosorbent assay and grouped into DGP high- and low-titer pools. The recognition pattern obtained after using both CD sera pools for immunoblotting did not reveal differences between control and MTG-treated bread protein extracts. Our results indicate that MTG treatment of wheat bread prepared with typical MTG concentrations used in standard bakery processes does not lead to immunodetectable amounts of CD immunotoxic deamidated gliadins.

Effects of Red Bean (Vigna angularis) Protein Isolates on Rheological Properties of Microbial Transglutaminase Mediated Pork Myofibrillar Protein Gels as Affected by Fractioning and Preheat Treatment.

Fractioning and/or preheating treatment on the rheological properties of myofibrillar protein (MP) gels induced by microbial transglutaminase (MTG) has been reported that they may improve the functional properties. However, the optimum condition was varied depending on the experimental factors. This study was to evaluate the effect of red bean protein isolate (RBPI) on the rheological properties of MP gels mediated by MTG as affected by modifications (fractioning: 7S-globulin of RBPI and/or preheat treatment (pre-heating; 95℃/30 min): pre-heating RBPI or pre-heating/7S-globulin). Cooking yields (CY, %) of MP gels was increased with RBPI (p<0.05), while 7S-globulin decreased the effect of RBPI (p<0.05); however, preheating treatments did not affect the CY (p>0.05). Gel strength of MP was decreased when RBPI or 7S-globulin added, while preheat treatments compensated for the negative effects of those in MP. This effect was entirely reversed by MTG treatment. Although the major band of RBPI disappeared, the preheated 7S globulin band was remained. In scanning electron microscopic (SEM) technique, the appearance of more cross-linked structures were observed when RBPI was prepared with preheating at 95℃ to improve the protein-protein interaction during gel setting of MP mixtures. Thus, the effects of RBPI and 7S-globulin as a substrate, and water and meat binder for MTG-mediated MP gels were confirmed to improve the rheological properties. However, preheat treatment of RBPI should be optimized.

Microbial transglutaminase treatment in pasta-production does not affect the immunoreactivity of gliadin with celiac disease patients' sera.

The effect of microbial transglutaminase (MTG)-treatment of pasta-dough on the immunoreactivity with celiac disease patient's sera has been investigated. Modification by MTG has been proven by determination of the MTG reaction product ε-(γ-glutamyl)lysine (3.63 μmol/g protein), which was not detectable in non-MTG-treated pasta. Antigenicity has been analyzed by immunoblotting and ELISA using gliadin-extracts from pasta and MTG-treated pasta. Immunoblotting showed that the antibody-population (antigliadin antibodies and antideamidated gliadin antibodies) of the sera is specific for every individual patient. Immunoblotting and ELISA showed that there is no difference in immunoreactivity of gliadin extracted from pasta and MTG-pasta. Recognition pattern and intensity in Western blot as well as antibody titer has also been identical even for sera with a high antideamidated gliadin antibody titer. These results indicate no difference between pasta-gliadin and MTG-pasta-gliadin and especially no increased deamidation in pasta-gliadin by MTG-treatment.

Oxidation-Induced Unfolding Facilitates Myosin Cross-Linking in Myofibrillar Protein by Microbial Transglutaminase

Myofibrillar protein from pork Longissimus muscle was oxidatively stressed for 2 and 24 h at 4 °C with mixed 10 μM FeCl(3)/100 μM ascorbic acid/1, 5, or 10 mM H(2)O(2) (which produces hydroxyl radicals) and then treated with microbial transglutaminase (MTG) (E:S = 1:20) for 2 h at 4 °C. Oxidation induced significant protein structural changes (P < 0.05) as evidenced by suppressed K-ATPase activity, elevated Ca-ATPase activity, increased carbonyl and disulfide contents, and reduced conformational stability, all in a H(2)O(2) dose-dependent manner. The structural alterations, notably with mild oxidation, led to stronger MTG catalysis. More substantial amine reductions (19.8-27.6%) at 1 mM H(2)O(2) occurred as compared to 11.6% in nonoxidized samples (P < 0.05) after MTG treatment. This coincided with more pronounced losses of myosin in oxidized samples (up to 33.2%) as compared to 21.1% in nonoxidized (P < 0.05), which was attributed to glutamine-lysine cross-linking as suggested by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

Effect of microbial transglutaminase treatment on thermal stability and pH-solubility of heat-shocked whey protein isolate

Whey protein isolate (WPI) dispersions (5% protein, pH 7.0) were subjected to heat-shock at 70 °C for 1, 5 and 10 min. The heat-shocked WPI dispersions were treated with microbial transglutaminase (MTGase) enzyme, and thermal properties and pH-solubility of the treated proteins were investigated. Heat-shocking of WPI for 10 min at 70 °C increased the thermal denaturation temperature (Td) of β-lactoglobulin in WPI by about 1.5 °C. MTGase treatment (30 h, 37 °C) of the heat-shocked WPI significantly increased the Td of β-lactoglobulin by about 6.3–7.3 °C when compared with heat-shocked only WPI at pH 7.0. The Td increased by about 13–15 °C following pH adjustment to 2.5; however, the Td of heat-shocked WPI was not substantially different from heat-shocked and MTGase-treated WPI at pH 2.5. Both the heat-shocked and the heat-shocked-MTGase-treated WPI exhibited U-shaped pH-solubility profiles with minimum solubility at pH 4.0–5.0. However, the extent of precipitation of MTGase-treated WPI samples at pH 4.0–5.0 was much greater than all heat-shocked and native WPI samples. The study revealed that while MTGase cross-linking significantly enhanced the thermal stability of β-lactoglobulin in heat-shocked WPI, it caused pronounced precipitation at pH 4.0–5.0 via decreasing the hydrophilic/hydrophobic ratio of the water-accessible protein surface.

Gelation properties of salt-extracted pea protein isolate catalyzed by microbial transglutaminase cross-linking

Gelation is a fundamental functional characteristic of plant proteins. In this paper, a salt-extracted pea protein isolate (PPI) was mixed with microbial transglutaminase (MTG) to produce gels and the gelation properties were studied. When the MTG level increased, the magnitude of both the G′ and G″ moduli also increased, which means the gel strength increased. A second order polynomial equation was used to describe the relationships between the G′, G″ modulus and TG level. It was found that with increased heating and cooling rate at the same MTG level, G′ and G″ tended to decrease, resulting in a weaker gel. This was attributed to the rearrangement time of pea protein molecules; slower heating and cooling rates enabled protein molecules to have more time to rearrange and therefore form a stronger gel. At the same MTG level, higher pea protein concentration resulted in higher G′ and G″ values and a power law relationship was found between G′ and pea protein concentration or G″ and pea protein concentration. Frequency sweep data of PPI show that the MTG treatment resulted in higher G′ values and lower tan delta values, indicative of a stronger more elastic gel. The minimum gelation concentration was found to be 3% (w/v) with 10 U MTG treatment, lower than 5.5% required when no MTG was present. When compared to PPI and soy protein isolate (SPI) with and without 10 U MTG treatment, the gel strength of PPI with MTG was stronger than that of SPI with MTG treatment, whereas the opposite was true without the MTG treatment. SDS-PAGE showed that at the same pea protein concentration, higher MTG level induced more cross-linking as fainter bands were seen on the gel and there was a shift in the relative intensities of the bands in the molecular weight range of 35–100 kDa.

Application of Microbial Transglutaminases in Anti-Crease Finish of Silk Fabric

In this paper, microbial transglutaminase (MTG) was applied to process silk fabric for improving its crease resistance under the prerequisite of maintaining other performances. Not only was the effect of MTG on silk fabric investigated through the Fourier transform infrared spectroscopy (FR), but analysis was also undertaken in the microcosmic structure of fibroin through the scanning electron microscope (SEM). Solo MTG treatment as well as compound treatments of MTG followed by hydrogen peroxide, protease and ultrasonic, all showed that MTG can improve the crease resistance of silk fabric. It also enhanced its tensile breaking strength or amended damage in the tensile breaking strength caused by pretreatments.Simultaneously, comparison with other treatments showed that compound treatment of MTG followed by ultrasonic exerted a better coordinated effect and conferred better performances, which made the wrinkle recovery angle (WRA) increase by 17.4% and tensile breaking strength improve by 11.2% respectively. At the same time, other performances were still maintained well.

Inconsistency in the Improvements of Gel Strength in Chicken and Pork Sausages Induced by Microbial Transglutaminase

This research investigated variation in the improvement of the texture of chicken and pork sausages induced by microbial transglutaminase (MTG). The extractability of myofibrillar proteins from these sausages as well as the e-(γ-glutamyl)lysine (G-L) content were also investigated. MTG treatment of sausages significantly increased the breaking strength values in both meat types, especially for samples incubated at 40°C. However, values of the breaking strength in both meat types were increased differently. The variation in protein extractability of samples incubated at 40°C for both meat types could lead to some consideration of the mechanisms and the high accessions of myosin heavy chain (MHC) to MTG. SDS-PAGE analysis showed significant changes in the density of the bands after adding MTG, especially for the pork samples in which the bands disappeared totally. The G-L content in the presence of MTG was double that in control samples of both meat types. This study suggests that the binding ability of myofibrillar proteins with MTG is strong. This leads us to suggest that MTG functions positively with different improvements in the texture of chicken and pork products that are treated mechanically, such as sausages. Variability in gel improvement level between chicken and pork sausages was observed; this resulted from the variation in meat proteins in response to MTG, as well as to the original glutamyl and lysine content.

Improvement of shrink-resistance and tensile strength of wool fabric treated with a novel microbial transglutaminase from Streptomyces hygroscopicus

In this study, a novel microbial transglutaminase (MTG) from Streptomyces hygroscopicus WSH03-13 was applied in the processing of wool fabrics. The results indicated that MTG treatment could improve felting properties and decrease tensile strength loss of wool fabrics. For the wool fabrics used in this study, MTG treatment following chemical and protease pretreatment led to a 2.32% of area shrinkage and about 16% recovery in tensile strength based on the samples without MTG treatment. Moreover, a traditional resin treatment was compared with the role of MTG. Although the tensile strength of wool fabrics treated by MTG was lower than that treated by resin treatment, the fabrics had similar anti-felting properties, and the chemical oxygen demand of wastewater was only half of the latter.

Effect of transglutaminase treatment on the glass transition of soy protein.

The effect of microbial transglutaminase (MTG) treatment on the glass transition temperature (T(g)) of two fractions which were isolated from a soy protein sample was studied. The T(g) of each fraction measured by differential scanning calorimetry was lowered by the MTG treatment, which generated cross-links in the samples, and this result agreed with the result of dynamic mechanical analysis. From the (1)H NMR measurement, the line width of the (1)H signal of the MTG-treated sample was observed to be greater than that of the MTG-nontreated sample at similar water content, which implied that there was relatively more immobilized water in the MTG-treated sample. The MTG treatment seemed to cause the increment in immobilized water, which might affect the T(g) of the soy protein sample.

Relationship between the glass transition of soy protein and molecular structure.

The change in molecular structure of the soy protein samples as a result of the microbial transglutaminase treatment was studied using solid-state (13)C NMR spectroscopy and circular dichroism (CD), and the relation to the glass transition temperature (T(g)) was examined. From NMR measurements, the structure of the local region of the C(alpha) methine was observed to change, and the region had relatively high mobility. From CD measurements, the structural change seemed to be caused by the change in the secondary structure (disintegration of the beta-structure). By comparison with the T(g) of another protein, the state of the secondary structure of a protein was suggested to be a key in determining its T(g).

Effects of hen egg yolk immunoglobulin in passive protection of rainbow trout against Yersinia ruckeri.

Anti-Yersinia ruckeri egg yolk immunoglobulin (IgY) was transferred to egg yolk after immunization of White Leghorn hens with formalin-killed whole cells of serovar 1 (RS1154) and serovar 2 (RS1153)Y. ruckeri and its lipopolysaccharide (LPS). The IgY was specific for its homologous LPS in western immunoblot, whereas some protein bands were commonly recognized, even by IgY from eggs of unimmunized hens. Purified LPS from both Y. ruckeri serovar types 1 and 2 had a very poor immunogenicity. The IgY activity was stable when processed into pellet form by a microbial transglutaminase treatment and showed a considerable resistance against acid pepsin for at least 2 h. Feeding specific anti-serovar 1 Y. ruckeri IgY to fish either before or after immersion infection produced marginal reductions in mortalities and in intestine infection. The same IgY did passively protect rainbow trout against infection when administered by intraperitoneal injection 4 h before an immersion challenge.

Glass Transition Temperature of Casein as Affected by Transglutaminase

ABSTRACT:The objective of this study was to determine glass transition temperature (Tg) of a‐casein with differential scanning calorimetry (DSC) and to study the effect of microbial transglutaminase (MTG) treatment on Tg of a‐casein. MTG‐treated a‐casein was fractionated following the MTG treatment. After adjusting water content, T measurements were made by DSC and the dependencies of Tg on the water content determined. Oscillation DSC (ODSC) was also used. The determination of Tg of MTG‐treated samples by conventional DSC was unreliable. However, by comparing with ODSC data, their T could be detected. The MTG treatment elevated the Tg of a‐casein, which maybe the reason for improved stability of dry daily products during storage.