The peptide deformylase (PDF) is a unique feature of prokaryotes and believed not to exist in eukaryotes up to even only a few years ago. PDF is essential for ribosomal protein translation by deformylating the initiating N-formylmethionine in prokaryotes. Since the initiating methionine of eukaryotes is not N-formylated, PDF inhibitors were considered as a possible antibacterial drug target. Many researchers have studied PDF and PDF inhibitors to overcome the drug resistance problem. Currently, there are 280 papers on the structure and function of bacterial PDF, and 103 papers on bacterial PDF inhibitors. Among them, LBM415 and BB-83698 are already under clinical study as novel antibiotics candidates.
However, PDF also exists in the mitochondria of eukaryotes including humans, and is crucial for cell growth according to recent papers. The first eukaryotic PDF was reported to be in the mitochondria and plastids of Arabidopsis thaliana. A bioinformatics study found many PDF-like genes in parasites, plants, and mammals. In 2004, Lee and coworkers reported that non-peptidyl bacterial PDF inhibitor actinonin suppresses the proliferation of cancer cells more than normal cells through a tumor-specific mitochondrial membrane depolarization and ATP depletion mechanism by interacting with HsPDF. It also could remove the N-formyl capping of model human mitochondrial peptides in prokaryotes. However, the function of HsPDF in vivo was unclear. The crystal structure of HsPDF was determined by EscobarAlvarez et al. and the mechanism of action was proposed. It is known that HsPDF processes formylated peptides derived from mitochondrial DNA-encoded proteins by a Co2+ dependent manner. The bacterial PDF and HsPDF have both similarities and differences in their structures. The complex structure of HsPDF and actinonin, a naturally occurring inhibitor derived from the Streptomyces species, shows an overall structural similarity to the complex structure of bacterial PDF and actinonin, but they have structural differences at the C-terminus.
In the previous study, we made novel bacterial PDF inhibitors, PMT analogs, which named after ProMediTech Ltd. In the progress of developing PMT analogs, we discovered about 600 kinds of compounds in our library. PMT analogs have potent antibacterial activity against Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis. These bacteria were known to cause respiratory tractassociated infections. Structural modification of PMT analogs makes these compounds more effective especially against Staphylococcus aureus. PMT analogs have a peptidomimetic structure like actinonin, but have stronger activity and better pharmacokinetic properties than actinonin.
Based on our previous studies, we predicted that PMT analogs could bind to HsPDF and inhibit the proliferation of cancer cells like actinonin. In order to prove our hypothesis, we carried out virtual screening of PMT analogs with GOLD software and DOCK6.5 from UCSF and selected 43 compounds from our library. The MTT assay was performed to test the antitumor activity for 10 highly scored compounds out of the 43 compounds. Among the 10 compounds, 4 analogs, PMT-172, PMT-173, PMT-199, and PMT-201, showed similar or better activity compared to actinonin. We prepared a HsPDF construct that has an enhanced expression level by codon optimization in E. coli. Then, 1H-CPMG-T2 filter NMR experiments were performed with purified protein, and it was confirmed that PMT-172 and PMT-199 could bind directly to HsPDF. The docking models between PMT-172, PMT-199 and HsPDF from the X-ray crystal structure (PDB 3G5K) with DOCK6.5 were obtained. It was identified that PMT-172 and PMT-199 have an N-alkyl amide and sulfonyl amide group, respectively, at the functional group R3 and R4. Understanding the structural characteristics of PMT-172 and PMT-199 might yield clues to design novel anticancer drug candidates that have better physical properties and binding affinities through modification of the functional groups.
Reference:
Kim, Won-Je, et al. “Bacterial peptide deformylase inhibitor PMT analogs inhibit cancer cell growth by interacting with human peptide deformylase.” Chinese science bulletin 59.32 (2014): 4274-4282.