Discovery of Two Brominated Oxindole Alkaloids as Staphylococcal DNA Gyrase and Pyruvate Kinase Inhibitors via Inverse Virtual Screening

Abstract

In the present study, a small marine-derived natural products library was assessed for antibacterial potential. Among 36 isolated compounds, a number of bis-indole derivatives exhibited growth-inhibitory activity towards Gram-positive strains (Bacillus subtilis and multidrug-resistant Staphylococcus aureus). 5- and 6-trisindoline (5-Tris and 6-Tris) were the most active derivatives (minimum inhibitory concentration, MIC, 4-8 µM) that were subsequently selected for anti-biofilm activity evaluation. Only 5-Tris was able to inhibit the staphylococcal biofilm formation starting at a 5 µM concentration. In order to investigate their possible molecular targets, both natural products were subjected to in silico inverse virtual screening. Among 20 target proteins, DNA gyrase and pyruvate kinase were the most likely to be involved in the observed antibacterial and anti-biofilm activities of both selected natural products. The in vitro validation and in silico binding mode studies revealed that 5-Tris could act as a dual enzyme inhibitor (IC₅₀ 11.4 ± 0.03 and 6.6 ± 0.05 µM, respectively), while 6-Tris was a low micromolar gyrase-B inhibitor (IC₅₀ 2.1 ± 0.08 µM), indicating that the bromine position plays a crucial role in the determination of the antibacterial lead compound inhibitory activity.

Keywords: MRSA; antibacterial; antibiofilm; gyrase-B; inverse virtual screening; marine natural products; pyruvate kinase; trisindoline.

Figure 1

Some examples of marine-derived indole compounds.

Figure 2

Outline of the procedure used in this study. SAR: structure–activity relationship.

Figure 3

Inhibition of biofilm formation and planktonic growth of Staphylococcus epidermidis RP62A by (A) 5-trisindoline (5-Tris) and (B) 6-trisindoline (6-Tris). OD: Optical Density.

Figure 4

Structure–activity relationship of the bioactive indole compounds. PK: pyruvate kinase; Gyr-B: gyrase subunit B.

Figure 5

Docking study of 5-Tris (A,B) and 6-Tris (C,D) within the active site of multidrug-resistant Staphylococcus aureus (MRSA) Gyr-B. The key binding interactions of the Gyr-B co-crystallized ligand [29] are shown in (E,F). The amino acid side chains are depicted in (A,C,E) for clarification.

Figure 6

Docking study of 5-Tris (A,B) and 6-Tris (C,D) within the active site of MRSA PK. The key binding interactions of the PK co-crystallized ligand [21] are shown in (E,F). The amino acid side chains are depicted in (A,C,E) for clarification.