The occurrence of multi-drug resistant Gram-negative pathogens compounded by a slow progress in discovering any new drugs has created an alarming situation worldwide. Bacteriophages and bacteriophage-derived peptidoglycan hydrolases (endolysins) represents a promising alternative for the treatment of multi-drug resistant Gram-negative and Gram-positive pathogens. In this study, Gp105 predicted to be a lysozyme murein hydrolase, from Enterobacter phage mypsh1140 has been identified and characterized through in silico analysis. The sequence analysis showed Gp105 to have T4-type_lysozyme like domain (IPR001165) and belongs to Glycoside hydrolase, family 24 (IPR002196). Further, the gene was cloned into pET28a vector, expressed in E. coli BL21 (DE3) and purified by Nickel affinity chromatography. In future, the purified protein will be utilized for its therapeutic potential by studying its antimicrobial properties against wide bacterial hosts and its ability to act in synergy with other antimicrobial agents.