**5.4 Nucleotide interaction**

AMPs show interaction with nucleotides like Adenosine triphosphate (ATP) [65]. In biofilms, one such crucial mechanism for AMPs is to interact with the

alarmone nucleotides (p) ppGpp which can be detected by a co-precipitation assay. This interaction results in ppGpp degradation in bacteria, blocking the stress reaction, which further results in the prevention of biofilms or removal of already formed biofilms [7].

#### **5.5 Other methods**

Several other methods include determining different types of interactions, for instance, the capacity of AMPs to interact with protein molecules to prevent the formation of biofilm [66]. The co-precipitation method can be used, to detect the capability of AMPs to interact with proteins. For example, the ribosomal protein binding activity of Bac71–35 was examined by measuring the activity of cosedimentation of ribosomes that have been purified with Bac71–35. After incubating *E. coli* 70S ribosomes with various doses of the peptide, the peptide bound to the ribosome was isolated using ultracentrifugation. Immunoblotting was used to validate the existence of Ba71–35 and ribosomal protein interaction in the ribosomal pellets [56].

Alternatively, the presence of peptide can be detected by labeling it with rhodamine whether the peptide is on the membrane surface of bacteria or inside the bacteria or on a solid attachment [66] or fluorescent dyes [57]. It is crucial to confirm that the label should not cause any interference with the activity and composition of peptides. Finally, several other kinds of interactions (for instance, lipid II or LPS) can be detected by the use of techniques such as Nuclear Magnetic Resonance (NMR) or surface plasmon resonance (SPR) [7].

#### **6. Conclusion**

Antimicrobial peptides are an essential component of innate immunity. They have the potential to be a viable alternative to antibiotics. It is critical to comprehend the MOA used by AMPs to kill bacteria to increase their development as therapeutics. The selectivity and activity of these peptides are influenced by a variety of parameters. Properties such as net charge, hydrophobicity, secondary structure, and amphipathicity are all important for function and are so interconnected that changing one attribute generally causes alterations in others.

The AMPs aggregate at the membrane surface following the hydrophobic and electrostatic attraction, forming self-aggregate on the bacterial membrane after they reach a particular concentration. The MOA of peptides can be broadly divided into two classes including direct killing and immunological regulation, wherein direct killing is further categorized into membrane targeting and non-membrane targeting. Different models have been proposed to explain the mechanism of interaction of peptides with membranes. Some biophysical techniques are used to determine their action mechanism whether these peptides disrupt microbial membrane or they target intracellular activities. Therefore, it is very crucial to know about the diverse biological features of AMPs for their clinical development.

*Antimicrobial Peptides: Mechanism of Action DOI: http://dx.doi.org/10.5772/intechopen.99190*
