Proteases are the major enzymes that have been used widely in health, food, and nutritional supplement fields. Rare actinobacteria are considered a new resource for proteolytic enzyme activity. Rare actinobacteria are used to be isolated from marine and have been studied under restricted limits. The strain used in this study was identified as Lentzea sp. which was successfully isolated from the soil at the Faculty of Science, Damietta, Egypt. It was subjected to gelatin liquefaction as primary screening for gelatinolytic activity. Based on this study, the best gelatinolytic (gelatinase) activity was noticed within 8 days at 37oC (17.59 U/ml ±0.06). The maximum activity was obtained at pH 9 (28.33 U/ml ±0.04) and agitation speed of 150 rpm (29.10 U/ml ±0.04). Upon carbon and nitrogen source optimization, the basal medium of combination between gelatin and peptone produced maximal gelatinase activity (29.12 U/ml ±0.06) and it was found that 10% sucrose was the best carbon source that produced maximum gelatinase activity at (30.17 U/ml ±0.05).