## 3 Antibacterial Peptides and Their Mechanism of Action
There, they can affect different enzymes and intermediates involved in vital processes. The inhibitors of the nucleic acid biosynthesis seem to have a high binding affinity for both DNA and RNA because they share with nucleic acid-binding enzymes or substrates, homologous fragments of their sequences; an interesting example is represented by DNA-binding protein histone H2A [73]. Other mechanisms use the inhibition of the enzymes involved in the DNA/RNA biosynthesis, like DNA topoisomerase I preventing DNA relaxation [74], RNA polymerase blocking the transcription [75] and gyrase impairing the supercoiling of DNA. [76] As a result, DNA/RNA degradation is induced and consequentially also cell death. There are several inhibitors of protein biosynthesis which alter the transcription and the translation but also the correct folding and the degradation of the protein. Usually, the AMPs that act on the protein biosynthesis target the ribosome subunits [77] but some others can interfere with the incorporation of histidine, uridine and thymidine [78; 79], the amino acid synthesis pathways [80], the release factors on the ribosome [81], the regulation of sigma factors [82], the nucleotide and coenzyme transport [80] and the degradation of DNA-replication-associated proteins [83]. Some peptides influence protein folding, in particular, DnaK, the major Hsp70 of the chaperone pathway in _Escherichia coli_, which has been seen as an optimal target to prevent the refolding of misfolded proteins [84]. Another approach is linked to the inhibition of matrix metalloproteases, essential enzymes in microbial cell growth and homeostasis, i.e., serine protease, trypsin-like protease, elastase and chymotrypsin [85; 86; 87]. There are also inhibitors of cell division that block DNA replication or the mechanisms essential for the repair of DNA damages, then resulting in the block of the cell cycle, in the impairment of the chromosome separation, in the failure of septation, in the alteration of mitochondrial activity and in a substantial change in the cell morphology with clearly visible blebbing and elongation towards a filamentous shape [88; 89]. Cell wall synthesis is another suitable target. Some AMPs act on lipid II by sequestrating it from the functional site [90; 91] or by binding D-Ala-D-Ala residues of its precursor preventing the addition of \(N\)-acetylglucosamine and \(N\)-acetylmuramic acid in the structure, hence the peptidoglycan elongation [92]. Other peptides have shown antimicrobial activity by activating cell wall-associated lytic enzymes, for example, some AMPs binding teichoic and teichuronic acids which otherwise are linked to amidases. The release of amidase stimulates premature autolysin activity and, consequently, cell lysis. Moreover, lipopolysaccharide-LPS are components of the membrane as well but, when released, are also well-known endotoxins able to raise an excessive and harmful pro-inflammatory response. AMPs that bind and neutralize LPS avoid the excessive stimulation of the immune system favoring a correct and balanced infection resolution [93]. Recently, the AMPs inhibitory activity on biofilm has been reported. Biofilm, consisting of an extracellular matrix of mainly polysaccharides, provides virulence, persistence and drug resistance to the microbial community [94; 95]. Anti-biofilm mechanisms, similar to the membrane-targeting ones, are also very diverse and sequence dependent. A database of biofilm-active peptides can be found online [[[http://www.baamps.it/](http://www.baamps.it/)]([http://www.baamps.it/](http://www.baamps.it/)), accessed on 10 November 2021]. AMPs could prevent biofilm formation by affecting cell attachment, or could act on preformed biofilm by disrupting the _quorum-sensing_, dispersing the cells within it, or affecting the expression of the related genes [96; 97]. Destabilization of matrix architecture impairing secretion or interaction between the matrix polymers has been also hypothesized [98].