Antimicrobial Activity of Diphenyl Pyridine Phosphine Gold(I)-thiolateCompounds and their Molecular Docking With Thioredoxin Reductase Enzyme

The spread of resistance to antimicrobial agents is currently considered as one of the major global health problems. Treatment-resistant hospital-acquired infections can cause an increase in mortality rate and treatment costs. Currently, Staphylococcus aureus, enterococci, Pseudomonas aeruginosa, Escherichia coli and Candida albicans are the main causes of hospital-acquired infections. Due to the prevalence of fungal and bacterial infections in recent years, efforts are being made to design and synthesize novel compounds that are effective against these organisms. Evaluation of the antimicrobial activity of the diphenyl pyridine phosphine gold(I)-thiolate compounds was first done by Robert Koch at the end of the 19th century, who showed that potassium dicyanidoaurate (I) with formula K[Au(CN)2], had antimicrobial activity against Mycobacterium tuberculosis. In the mid-20th century, gold(I) complexes were used clinically as anti-inflammatory agents for the treatment of various rheumatic diseases, including psoriatic arthritis, juvenile arthritis, and palindromic rheumatism. Recent studies have shown that gold(I) compounds are promising candidates for making antimicrobial drugs. The interest in gold-based drugs is increasing. Inhibition of the thioredoxin reductase (TrxR) enzyme is the most important biological target for antimicrobial gold(I) compounds.

In this study, five diphenyl pyridine phosphine gold(I)-thiolate compounds underwent biological evaluation. All compounds contained the diphenyl-pyridinephosphine ligand. They were attached to chlorine in the Au1 complex and to thiolate ligands (2-mercaptopyridine, 2-mercaptopyrimidine, 2-mercaptobenzothiazole and 2-mercapto-dihydro-thiazole) in Au2- to Au5 complexes. Bacteria and yeasts were stored as glycerol stocks at -80 °C. Before each experimental, they were gently placed on Mueller-Hinton plates. Colonies of new prepared plates were incubated overnight at 37 °C in 5 mL of media. The overnight culture medium was diluted to OD600<0.01 in 2 mL of new prepared media in sterile culture tubes. The compounds (auranofin and synthesized gold compounds) were dissolved in dimethyl sulfoxide (DMSO) and a concentration of 1 mg/mL was prepared from them. Then, they were diluted at specific concentrations in cell culture tubes. Control tubes for each cell type were incubated with an equivalent concentration of DMSO. The 200-μL cultures were transferred to a 96-well plate. The plates were incubated at the appropriate temperature and were shaken in a microplate reader (ELx808, Biotek Co.). Growth measurements (OD600) were performed automatically every hour for 24 hours. The minimum inhibitory concentration (MIC) was determined as the concentration at which there was a negligible increase in the OD600 value from the initial reading after 24 hours.Molecular docking studies of these compounds were performed on TrxR enzyme using AutoDock 4.2. In this regard, the best X-ray crystallographic structure of a TxrR enzyme with PDB code of 4CBQ was extracted from the protein data bank. The ranking was done only based on the ligand-receptor binding free energy data from the final AutoDock docking log file (dlg). MOE 2014 and ADT software were used to analyze the interaction of compounds with the receptor.

The results from analyzing the antibacterial effects of the study compounds on standard and clinical samples of Gram-positive and Gram-negative bacteria are shown in Table 1. The antibacterial activity of all compounds were compared with the antibacterial activities of auranofin (an anti-rheumatoid gold-containing drug) and ciprofloxacin. These compounds had the MIC values ranged from 3 to 100 μg/mL. The most active compound (Au3) had an MIC of 3.89, 3.15, 4.36, 5.44, 6.13, and 8.37 μg/mL against P. aeruginosa, E. coli, S. aureus, B. subtilis, C. albicans and S. cerevisiae, respectively. Molecular docking was performed to find out their binding site and binding mode to the TrxR. In evaluating the effects of antifungal and yeast activity of gold complexes on C. albicans and S. cerevisiae, the highest antifungal activity observed was related to the Au3 compound.The structure-activity relationship of gold(I) compounds showed that the absence of thiolate ligand significantly reduced the antimicrobial activity. The presence of aromatic thiolate ligand significantly increased the antimicrobial activity.The binding of these compounds to the amino acids of the active site of the TrxR enzyme was also investigated. As shown in Figure 1, the Au2 compound formed a weak hydrogen bonding with amino acid Thr269 through sulfur attached to its pyridine ring. The binding of gold atom to amino acid Cys286, which is one of the most important binding compounds of TrxR inhibitors, was also observed. 

In this study, the antimicrobial effects of five diphenyl pyridine phosphine gold(I)-thiolate compounds on two types of gram-negative bacteria (P. aeruginosa and E. coli), two types of gram-positive bacteria (S. aureus and B. subtilis), one type of fungus (C. albicans) and one type of yeast (S. cerevisiae) were evaluated. These compounds showed good antimicrobial activity and had high potential in inhibiting the growth of microbial cells. The best compound complex was Au3. These compounds act much better especially on gram-negative and yeast strains compared to auranofin as antirheumatic drug. These compounds, especially the Au3, are potentially valuable for drug development and can be a promising antimicrobial agent.