Further work on a thermophilic rennet synthesized by a thermophilic actinomycete is reported. It was produced by growth in fermentor of 50‐or‐200‐L volume and was purified by membrane filtration of a cell‐free supernatant and then molecular filtration and ion exchange chromatography. Its stability under various conditions was determined: The enzyme is a true rennet requiring calcium ions for activity. Experiment shows that it resembles neither pepsin, trypsin, nor chymotrypsin. It is freeze‐labile with a molecular weight of 9700 from amino acid composition. When skim milk powder was added to the growth medium, a rennet with different properties was obtained.