Stabilization Study Of Microbial Transglutaminase

Transglutaminase （protein-glutamine-γ-glutamytransferase, EC2.3.2.13, TGase） is a kind of acyltransferase that can catalyze acyl transfer reactions introducing covalent cross-links between proteins as well as peptides and various primary amines. These reactions can change the function properties of protein, reform structures and increase nutrition value. Transglutaminase has been found in animal, plant tissues and microorganisms. Commercial transglutaminase mainly comes from animal and microorganism. The complicated separation and purification procedure results in an extremely high price for the animal transglutaminase, which limits the wide application in food processing, while microbial transglutaminase produced by fermentation has more extensively application for its lower price and Ca²⁺-independent.The temperature is a main effect factor of the stability of MTG. for losing activity easily in high temperature. The influences of different carbohydrates, salts, protein and peptides to enzyme activity and the thermal stability of MTG were studied in this paper. The thermal stability of MTG was judged by calculating the residual enzyme activity after heated at specified temperature. The main results were as follows:1. The addition of sodium chloride or potassium chloride had only a slight influence on the activity of TGase. The addition of monovalent ions in low concentration decreased the thermal stability ,while the addition of monovalent ions in higher concentrations（exceed 10%） increased the thermal stability of TGase and a higher salt concentration lead to higher thermal stability. The residual enzyme activity increased 28.61% and 32.10% when 20% of NaCl and KC1 were added in the solution, respectively. The addition of bivalent ions reduced the enzymatic activity and thermal stability and a higher salt concentration lead to lower enzyme activity.2. The addition of glucose, sucrose, trehalose, lactose and maltitol reduced the activity of TGase, a higher concentration lead to lower enzyme activtity. The activity of TGase decreased 5.77 %when 10 % of lactose was added and the activities decreased 8.65 %,9.08 %,10.51 %, 8.74 % when 20% of glucose, sucrose, trehalose and maltitol were added, respectively. But those carbohydrates can increase the thermal stability. The higher concentration of carbohydrates lead to higher residual enzyme activity after TGase was heated at specified temperature. The residual enzyme activity increased 14.5% by 10% of lactose; The residual enzyme activity increased 28.46%, 27.52%, 26.69% and 26.47% with 20% of glucose, sucrose, trehalose and maltitol, respectively.3. Different kinds of polysaccharide had different influence on the thermal stability of TGase. The addition of xanthan gum and gum arabic had no influence on the TGase activity, while pectin decreased the enzyme activity. Xanthan gum has a lower solubility, so the addition of xanthan gum has no influence on the thermal stability. The addition of low concentration gum arabic （2.5 %） slightly improved the thermal stability of TGase, while the addition of higher concentration gum arabic （5 %, 7.5 % and 10 %） decreased the thermal stability. A higher addition concertration lead to lower thermal stability. In general, the addition of polysaccharides has no positive effect to activity of TGase.4. Different kinds of polypeptide and protein had different influence on the thermal stability of TGase. The addition of WGPH increased the thermal stability of TGase obviously, 1 % of WGPH can enhanced the remained enzyme activity at 5.87 %. The addition of peptone, HVP and BSA had no influence on the thermal stability ,while the addition of low molecular weight soybean oligo-peptide decreased thermal stability. A higher WGPH concentration leaded to higher thermal stability of TGase. The addition of WGPH had slightly extended the primary enzyme activity.5. Wheat gluten partial hydrolyzates with different hydrolyzed methods had different influence on the thermal stability of TGase. WGPH hydrolyzed by HCl inhibited the enzyme activity obviously and decreased the thermal stability of TGase, for its strongly hydrolyzed condition. WGPH hydrolyzed by enzyme increased the thermal stability of TGase, but the effect decreased with the increase of hydrolysis time and DH increasing. The addition of WGPH with high DH even decreased the thermal stability of TGase.6. WGPH hydrolyzed by enzyme had reducing capacity and anti-oxidized capacity. Wheat gluten did not have reducing capacity. The reducing capacity increased during the process of hydrolysis, but the reducing capacity keep invariable finally.7. WGPH as protective agent can prevent the enzymatic activity loss and improve recovery rate during lyophillization. The lyophillization recovery rate was 99.3% with 6% of WGPH, higher than control of 10.5%. The protective effect is obviously.