In the modern-day world, antibiotic resistance is one of the most serious threats to both human health and food security. Finding new ways to prevent and overcome the formation of pathogen resistance to antibiotics is an extremely important and urgent task in modern medical science. All bacteria, except myco-plasmas, have cell walls in which various enzymes, receptors, transporters, channels, and antigens are located. This review is devoted to describing the structure of the major elements of bacterial cell walls and the enzymes involved in their biosynthesis, and is used as molecular targets for screening and selection of new effective antibiotics. Special attention has been paid to methods for studying the functional activity and inhibition of these targets. In addition, this review describes the functional characteristics of pore-forming proteins in the outer membrane of Gram-negative bacteria and the molecular mechanisms of antibiotic penetration through porin channels. Analysis of the structure and functional features of the targets of different classes of antibiotics is the basis for developing new strategies to overcome bacterial resistance.