Protegrin-5

We cloned the genes of three protegrins, a family of cathelin-associated antimicrobial peptides originally isolated from porcine leukocytes. Each gene comprised 4 exons and 3 introns, wherein Exon I encoded the signal sequence and the first 37 amino acids of cathelin, Exons II and III contained 36 and 24 additional cathelin residues and Exon IV contained the final two cathelin residues followed by the protegrin sequence. This quadripartite gene structure helps explain how structurally diverse antimicrobial peptides can be expressed on common, cathelin-containing precursors. Southern blot probed with an oligonucleotide specific for protegrin genes suggested that several identical or nearly identical protegrin genes were densely clustered in the pig chromosome.

We tested 20 protegrins against Chlamydia trachomatis serovar L2 (L2/434/Bu). Five of the protegrins had native structures; the others included nonamidated, enantiomeric, and truncated variants and peptides with <2 disulfide bonds. Antichlamydial activity resided principally in residues 5 to 15 of native protegrin PG-1, and optimal activity required both intramolecular disulfide bonds.

Protegrin pore formation is believed to occur in a stepwise fashion that begins with a nonspecific peptide interaction with the negatively charged bacterial cell walls via hydrophobic and positively charged amphipathic surfaces. There are five known nature protegrins (PG1-PG5), and early studies of PG-1 (PDB ID:1PG1) shown that it could form antiparallel dimer in membrane mimicking environment which could be a first step for further oligomeric membrane pore formation. Later, we solved PG-2 (PDB ID:2MUH) and PG-3 (PDB ID:2MZ6) structures in the same environment and for PG-3 observed a strong dαα NOE effects between residues R18 and F12, V14, and V16. These "inconsistent" with monomer structure NOEs appears due to formation of an additional antiparallel β-sheet between two monomers. It was also suggested that there is a possible association of protegrins dimers to form octameric or decameric β-barrels in an oligomer state. In order to investigate a more detailed oligomerization process of protegrins, in the present article we report the monomer (PDB ID: 2NC7) and octamer pore structures of the protegrin-5 (PG-5) in the presence of DPC micelles studied by solution NMR spectroscopy. In contrast to PG-1, PG-2, and PG-3 studies, for PG-5 we observed not only dimer NOEs but also several additional NOEs between side chains, which allows us to calculate an octamer pore structure of PG-5 that was in good agreement with previous AFM and PMF data.