Background and objective: Through ceating crosslink in proteins, and strengthening the protein-based food structures, MTGase can positively affect the serum holding capacity and gel firmness properties of foods. The purpose of this study is to study the effect of the addition of MTGase on the physicochemical and sensory properties of spinach yogurt.Materials and methods: In this study, the effect of using MTGase (0.1, 0.2 and 0.3 g L-1) was examined as a substitute for part of the milk protein concentrate in spinach yoghurt. The effect of enzyme on selected physicochemical (e.g. pH, titratable acidity, syneresis and viscosity) and organoleptic properties (texture, flavor, odor and overall acceptability) of spinach yoghurt was examined. Evaluations were performed at 0, 5, 10 and 15 days of storage at 4oC. Statistical analyzes were performed with SPSS 20 software. Average treatments were compared using one-way analysis.Results: In general, addition of different concentrations of MTGase caused no significant changes on the pH and acidity of yoghurt samples. However, it increased the viscosity and decreased the syneresis. It was shown that MTGase at concentration of 0.1 g L-1 could be considered as optimal concentration. Although higher concentrations could produce better properties, they are not economic because fewer concentration of enzyme was able to create a sample as similar to the control.Conclusion: The results showed that the MTGase is an acceptable substitute for milk protein concentrate in spinach stirred yoghurt.