A light yellow, whitish and greyish pigment, proteolytic actinomycetes strain was isolated from marine soil samples collected form the coastal area of Chennai, India. In this study 114 isolates of actinomycetes were isolated and identified by morphological studies. The isolate grows to produce circular colonies that had yellow, white and greyish substrate mycelium on starch casein agar. Its proteolytic activity was indicated by growth and clear zone appearance on casein skim milk agar plate method. From the total 114 isolates 8 showed proteolytic activity; among them one strain MB22 which showed higher protease activity with lower biomass and it was taken for further investigation. The growth conditions of protease production by identified actinomycetes were optimized with different fermentation period, pH, Temperature, carbon source, nitrogen source. Among the different carbon source, sucrose was found to be best source. Beef extract was found to be the optimum nitrogen source for protease enzyme production by the actinomycetes. The best source was found to be pH-4, for temperature 46ËšC. The amount of protein was estimated by Bradford method and the molecular weight was determined by Sodium Dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE).