The outer membrane (OM) of Gram-negative bacteria acts as a selective barrier that significantly influences antibiotic susceptibility. Antibiotics can enter the cell through two pathways: lipid-mediated for hydrophobic drugs and general diffusion porins for hydrophilic drugs. The OM's lipid and protein composition greatly affects antibiotic sensitivity, and modifications to these components are common in drug-resistant strains. This review discusses the molecular mechanisms of antibiotic permeation through the OM and the strategies bacteria use to resist antibiotics by altering these pathways. The OM is an asymmetric bilayer composed of phospholipids and lipopolysaccharides (LPS), with LPS playing a key role in forming a hydrophobic barrier. The LPS structure includes lipid A, a core oligosaccharide, and the O-antigen. Modifications to LPS, such as the addition of 4-aminoarabinose and phosphoethanolamine, reduce its negative charge and increase resistance to cationic agents like polymyxin B. These modifications are regulated by the PmrA/PmrB and PhoP/PhoQ two-component systems. Porins, such as OmpF, OmpC, and OprF, are essential for the diffusion of hydrophilic antibiotics. These porins have a β-barrel structure with a constriction zone that determines their size exclusion limit. The function of porins can be modulated by factors such as pH, voltage, and polyamines, which can lead to closure or inactivation of the pores. Mutations in porin proteins can reduce antibiotic permeability, contributing to resistance. Antibiotic resistance in Gram-negative bacteria can arise from alterations in porin expression or function, leading to reduced permeability. For example, the loss or functional change of porins in strains like E. coli, P. aeruginosa, and N. gonorrhoeae results in increased antibiotic resistance. Additionally, the presence of specific porins, such as OprD in P. aeruginosa, allows for the uptake of certain antibiotics despite reduced porin expression. The review highlights the importance of understanding the OM's lipid and porin properties in developing strategies to combat antibiotic resistance. By elucidating the molecular mechanisms of antibiotic permeation and resistance, researchers can design more effective therapies or new antibiotics targeting these surface-exposed entities.The outer membrane (OM) of Gram-negative bacteria acts as a selective barrier that significantly influences antibiotic susceptibility. Antibiotics can enter the cell through two pathways: lipid-mediated for hydrophobic drugs and general diffusion porins for hydrophilic drugs. The OM's lipid and protein composition greatly affects antibiotic sensitivity, and modifications to these components are common in drug-resistant strains. This review discusses the molecular mechanisms of antibiotic permeation through the OM and the strategies bacteria use to resist antibiotics by altering these pathways. The OM is an asymmetric bilayer composed of phospholipids and lipopolysaccharides (LPS), with LPS playing a key role in forming a hydrophobic barrier. The LPS structure includes lipid A, a core oligosaccharide, and the O-antigen. Modifications to LPS, such as the addition of 4-aminoarabinose and phosphoethanolamine, reduce its negative charge and increase resistance to cationic agents like polymyxin B. These modifications are regulated by the PmrA/PmrB and PhoP/PhoQ two-component systems. Porins, such as OmpF, OmpC, and OprF, are essential for the diffusion of hydrophilic antibiotics. These porins have a β-barrel structure with a constriction zone that determines their size exclusion limit. The function of porins can be modulated by factors such as pH, voltage, and polyamines, which can lead to closure or inactivation of the pores. Mutations in porin proteins can reduce antibiotic permeability, contributing to resistance. Antibiotic resistance in Gram-negative bacteria can arise from alterations in porin expression or function, leading to reduced permeability. For example, the loss or functional change of porins in strains like E. coli, P. aeruginosa, and N. gonorrhoeae results in increased antibiotic resistance. Additionally, the presence of specific porins, such as OprD in P. aeruginosa, allows for the uptake of certain antibiotics despite reduced porin expression. The review highlights the importance of understanding the OM's lipid and porin properties in developing strategies to combat antibiotic resistance. By elucidating the molecular mechanisms of antibiotic permeation and resistance, researchers can design more effective therapies or new antibiotics targeting these surface-exposed entities.