PACAP sequence modifications modulate the peptide antimicrobial activity against bacterial pathogens affecting aquaculture.

The global aquaculture industry has significant losses each year due to disease outbreaks. Antibiotics are one of the common methods to treat fish infections, but prolonged use can lead to the emergence of resistant strains. Aeromonas spp. Infections are a common and problematic disease in fish, and members of this genera can produce antibiotic resistant strains. Antimicrobial peptides (AMPs) have emerged as an alternative method to treat and prevent infections and pituitary adenylate cyclase activating polypeptide (PACAP) is a prominent member of this family. The objective of this research was to study PACAP's direct antimicrobial activity and its toxicity in fish cells. Four synthetic variants of the natural PACAP from Clarias gariepinus were tested in addition to the natural variant. The experimental results show a different antimicrobial activity against A. salmonicida and A. hydrophila of each PACAP variant, and for the first time show dependence on the culture broth used. Furthermore, the results suggest that the underlying mechanism of PACAP antimicrobial activity includes a bacterial membrane permeabilizing effect, classifying PACAP as a membrane disruptive AMP. This study also demonstrated that the five PACAP variants evaluated showed low toxicity in vitro, at concentrations relevant for in vivo applications. Therefore, PACAP could be a promising alternative to antibiotics in the aquaculture sector.

Bestatin and bacitracin inhibit porcine kidney cortex dipeptidyl peptidase IV activity and reduce human melanoma MeWo cell viability.

Bestatin and bacitracin are inhibitors of metallo aminopeptidases and bacterial proteases. However, their effects on other human peptidases, like dipeptidyl peptidase IV (DPP-IV, EC 3.4.14.5) are not established. Inhibitors of DPP-IV activity are used for treating type 2 diabetes mellitus, cancers and immune system diseases. Bacitracin and bestatin inhibited porcine membrane-bound DPP-IV (pDPP-IV) activity. Mechanisms were different, i.e. non-competitive with α > 1 (α = 3.9) and Ki value of 75 µM for bestatin, and competitive with Ki value of 630 µM for bacitracin. The binding mode in the tertiary complex enzyme:substrate:bestatin suggested the structural basis of the inhibitory effect and that bestatin is potentially selective for DPP-IV, ineffective vs. S9 family members dipeptidyl peptidase 8/9 and fibroblast activation protein. In the human melanoma MeWo cell line, bestatin and bacitracin inhibited aminopeptidase N (APN) and DPP-IV activities, reduced cell viability and increased DNA fragmentation, suggesting induction of apoptosis. Since bacitracin and bestatin are already marketed drugs, studying in depth the molecular mechanisms underlying their effects on melanoma cells is warranted. Additionally, bestatin emerges as a new lead compound for the development of DPP-IV inhibitors, and a promising dual APN/DPP-IV inhibitor for the treatment of pathologies in which both enzymes are upregulated.