Industrial enzyme applications.

Transglutaminases (Tgases) are a widely distributed group of enzymes that catalyse the post-translational modification of proteins by the formation of isopeptide bonds. This occurs either through protein cross-linking via epsilon-(gamma-glutamyl)lysine bonds or through incorporation of primary amines at selected peptide-bound glutamine residues. The cross-linked products, often of high molecular mass, are highly resistant to mechanical challenge and proteolytic degradation, and their accumulation is found in a number of tissues and processes where such properties are important, including skin, hair, blood clotting and wound healing. However, deregulation of enzyme activity generally associated with major disruptions in cellular homoeostatic mechanisms has resulted in these enzymes contributing to a number of human diseases, including chronic neurodegeneration, neoplastic diseases, autoimmune diseases, diseases involving progressive tissue fibrosis and diseases related to the epidermis of the skin. In the present review we detail the structural and regulatory features important in mammalian Tgases, with particular focus on the ubiquitous type 2 tissue enzyme. Physiological roles and substrates are discussed with a view to increasing and understanding the pathogenesis of the diseases associated with transglutaminases. Moreover the ability of these enzymes to modify proteins and act as biological glues has not gone unnoticed by the commercial sector. As a consequence, we have included some of the present and future biotechnological applications of this increasingly important group of enzymes.

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Transglutaminases: Nature’s biological glues

Some characteristics and applications of a transglutaminase, derived from a variant of Streptoverticillium mobaraense, namely microbial transglutaminase (MTGase), are considered MTGase, mass-produced at low cost by fermentation, catalyses the crosslinking of most food proteins through the formation of an e-(γ-glutamyl)lysine bond, in the same way as well-known mammalian enzymes However, MTGase is remarkable because it is calcium independent and its molecular weight is smaller than that of other known enzymes The results of many studies suggest that MTGase, as well as other transglutaminases, has many potential applications in food processing and in other areas

Transglutaminase and its use for food processing

Biocatalytic potential of microorganisms have been employed for centuries to produce bread, wine, vinegar and other common products without understanding the biochemical basis of their ingredients. Microbial enzymes have gained interest for their widespread uses in industries and medicine owing to their stability, catalytic activity, and ease of production and optimization than plant and animal enzymes. The use of enzymes in various industries (e.g., food, agriculture, chemicals, and pharmaceuticals) is increasing rapidly due to reduced processing time, low energy input, cost effectiveness, nontoxic and eco-friendly characteristics. Microbial enzymes are capable of degrading toxic chemical compounds of industrial and domestic wastes (phenolic compounds, nitriles, amines etc.) either via degradation or conversion. Here in this review, we highlight and discuss current technical and scientific involvement of microorganisms in enzyme production and their present status in worldwide enzyme market.

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Microbial enzymes: industrial progress in 21st century

The production of poly-(γ-glutamic acid) from microorganisms and its various applications

Transglutaminase is an enzyme that forms crosslinks between protein molecules. This crosslinkage has unique effects on protein properties, gelation capability, thermal stability, water-holding capacity, etc. A transglutaminase has been isolated from Streptoverticillium sp., and its practical use in the food industry realized. Transglutaminase is now widely used in seafood, surimi products, meat products, noodles/pasta, dairy products, baked goods, and so on. It has great potential to improve the firmness, elasticity, viscosity, heat stability, and water-holding capacity of prepared foods through the mild enzyme reaction. The overall applications of transglutaminase in the food industry are reviewed.

Transglutaminase: its utilization in the food industry

To produce restructured meat, microbial transglutaminase (MTGase) was evaluated for its ability to introduce covalent crosslinks between protein molecules. Pork muscle cubes were mixed with MTGase and held at 5°C for 2 hr for the enzyme reaction. Restructured products were analyzed for binding strength without cooking. MTGase treatment resulted in effective binding of meat pieces provided there was addition of salt (NaCI). To ensure proper binding without NaCI, some food proteins with MTGase were also investigated. Meat cubes in combination with MTGase and sodium caseinate showed acceptable bind, and sodium caseinate appeared to be a superior substrate for the crosslinking to meat proteins than soy protein, whey protein, or gelatin. These results suggest a useful method for producing restructured meat which can be distributed in the raw, chilled state.

Production of Restructured Meat using Microbial Transglutaminase without Salt or Cooking

To determine bioavailability, expressed as the protein efficiency ratio (PER) and biological value (BV) in rats, of the epsilon-(gamma-glutamyl)lysine [epsilon-(gamma-Glu)Lys] moiety in crosslinked proteins, we prepared heavily crosslinked [21.5 /micromol epsilon-(gamma-Glu)Lys/g casein] and intermediately crosslinked [13.6 micromol epsilon-(gamma-Glu)Lys/g casein] casein, using microbial transglutaminase. In Experiment 1, rats were assigned to one of four diets (heavily or intermediately crosslinked caseins, intact casein or non-protein diet) for 4 wk to evaluate the bioavailability of the epsilon-(gamma-Glu)Lys moiety in crosslinked casein as the sole source of dietary protein. Rats that were fed intact casein and the two crosslinked caseins had similar growth rates, PER, and BV, indicating that crosslinked caseins supported the growth of rats similarly to the intact casein. In Experiment 2, heavily crosslinked casein was added to wheat gluten-based diets in concentrations of 20 and 40 g/kg diet to evaluate the bioavailability of lysine in the epsilon-(gamma-Glu)Lys moiety of the casein as a lysine supplement for lysine-poor gluten. One of six diets (heavily crosslinked or intact casein diets in the two concentrations, gluten diet, or non-protein diet) was fed to rats for 4 wk. No significant differences in food intake, body weight gain, PER or BV were observed among rats fed the intact or crosslinked casein diets at either 2 or 4 g/100 g casein. These results suggest that the epsilon-(gamma-Glu)Lys moiety in crosslinked caseins are absorbed and therefore supplement the gluten. HPLC analysis of urine and feces of rats fed the crosslinked caseins actually confirmed that approximately 99% of the epsilon-(gamma-Glu)Lys moiety was absorbed in the body.

The ε-(γ-Glutamyl)lysine Moiety in Crosslinked Casein Is an Available Source of Lysine for Rats

To provide a composition containing a mineral absorption accelerator which accelerates the absorption of minerals. It contains minerals and degradated products of poly-γ-glutamic acid as a mineral absorption accelerator and may be used in foods in a variety of forms, including a beverage, gel, solid or powder form.

High-absorption mineral-containing composition and foods

An composition containing transglutaminase and collagen. The composition can be used to prepare bound food, preferably without the use of allergy-causing casein proteins.

Chiya Kuraishi

Papers

Transglutaminase: its utilization in the food industry

Production of Restructured Meat using Microbial Transglutaminase without Salt or Cooking

The ε-(γ-Glutamyl)lysine Moiety in Crosslinked Casein Is an Available Source of Lysine for Rats

High-absorption mineral-containing composition and foods

Enzyme preparation for use in the binding of food materials and process for producing bound food