We here characterize five globular endolysins, encoded by a set of Gram-negative infecting bacteriophages: BcepC6gp22 (Burkholderia cepacia phage BcepC6B), P2gp09 (Escherichia coli phage P2), PsP3gp10 (Salmonella enterica phage PsP3), K11gp3.5 and KP32gp15 (Klebsiella pneumoniae phages K11 and KP32, respectively). In silico, BcepC6gp22, P2gp10 and PsP3gp10 are predicted to possess lytic transglycosylase activity, whereas K11gp3.5 and KP32gp15 have putative amidase activity. All five endolysins show muralytic activity on the peptidoglycan of several Gram-negative bacterial species. In vitro, Pseudomonas aeruginosa PAO1 is clearly sensitive for the antibacterial action of the five endolysins in the presence of the outer membrane permeabilizer EDTA: reductions are ranging from 1.89 to 3.08 log units dependent on the endolysin. The predicted transglycosylases BcepC6gp22, P2gp10 and PsP3gp10 have a substantially higher muralytic and in vitro antibacterial activity compared to the predicted amidases K11gp3.5 and KP32gp15, highlighting the impact of the catalytic specificity on endolysin activity. Furthermore, initial data exclude the synergistic lethal effect of a combination of the predicted transglycosylase PsP3gp10 and the predicted amidase K11gp3.5 on PAO1. As these globular endolysins show a lower enzymatic and antibacterial activity, in comparison to modular endolysins, we suggest that the latter should be favored for antibacterial applications.