Membrane-interaction and assembly of a leucine zipper peptide (LZP), and its single (SASA) and double (DASA) alanine-substituted analog onto mammalian, hRBCs and 3T3 cells and bacteria, Escherichia coli and Staphylococcus aureus were studied as a model system to understand the plausible role of assembly on their contrasting cytotoxic but similar bactericidal activities. Peptides' ability to depolarize and damage the membrane organization of hRBC and 3T3 cells decreased from LZP to SASA and to DASA which may be related to their decrease in assembly onto these mammalian live cells and oligomerization states in the presence of these cell membranes or zwitterionic PC/Chol lipid vesicles. However, LZP and its analogs showed appreciable similarities in damaging or depolarizing the E. coli or S. aureus cells, which further matched with their comparable assembly and oligomerization either onto these live cells or the cell membranes or in the presence of negatively charged PC/PG lipid vesicles.