Transglutaminase was purified to homogeneity from leaves of soybean (Glycine max). The molecular weight of the enzyme estimated by gel filtration and sodium dodecyl sulfate polyacrylamide gel electrophoresis was 80,000 daltons. This purified enzyme catalyzed the incorporation of [14C]-putrescine into N,N'-dimethylcasein as a protein substrate. With N,N'-dimethylcasein, the Km values for putrescine, spermidine and spermine were 109, 42 and 69 microM, respectively. The optimum pH and temperature for the enzyme reaction were 7.6 and 37 degrees C. Ca2+ was not an absolute requirement for enzyme activity unlike animal transglutaminases. The enzyme was activated by dithiothreitol, but inhibited by GTP. With molecular weight of this enzyme, this inhibition of enzyme activity by GTP indicates that this enzyme is very similar to known tissue transglutaminases in animals.