Na, K-ATPase as a Biological Target for Gold(III) Complexes: A Theoretical and Experimental Approach.

BACKGROUND: Gold-based complexes represent a new class of potential metallodrugs. Although their action mechanism is not entirely understood, it was shown that gold complexes inhibit some enzymes' activities. Among them, Na,K-ATPase is emerging as an essential target for various anticancer drugs. The functionalization of nanoparticles by gold(III) complexes could facilitate their delivery into the cells and enable the following of their distribution in the target tissues. OBJECTIVE: The paper presents an overview of Na,K-ATPase interaction with representative and structurally related cytotoxic gold(III) complexes. The results obtained by the employment of theoretical methods (DFT and docking studies) combined with the experimental approach involving a variety of nanotechnology-base techniques (UV/Vis, Raman and fluorescence spectroscopy, CD, AFM, DLS) are discussed. Detailed information was obtained on the enzyme's conformational and structural changes upon binding the gold(III) complexes. The experimentally determined reaction parameters (constants of dissociation and the reaction stoichiometry) were predicted theoretically. CONCLUSION: The presented results offer further support to the view that Na,K-ATPase may be a relevant biomolecular target for cytotoxic gold(III) compounds of medicinal interest.

Conjugates of Gold Nanoparticles and Antitumor Gold(III) Complexes as a Tool for Their AFM and SERS Detection in Biological Tissue.

Citrate-capped gold nanoparticles (AuNPs) were functionalized with three distinct antitumor gold(III) complexes, e.g., [Au(N,N)(OH)2][PF6], where (N,N)=2,2'-bipyridine; [Au(C,N)(AcO)2], where (C,N)=deprotonated 6-(1,1-dimethylbenzyl)-pyridine; [Au(C,N,N)(OH)][PF6], where (C,N,N)=deprotonated 6-(1,1-dimethylbenzyl)-2,2'-bipyridine, to assess the chance of tracking their subcellular distribution by atomic force microscopy (AFM), and surface enhanced Raman spectroscopy (SERS) techniques. An extensive physicochemical characterization of the formed conjugates was, thus, carried out by applying a variety of methods (density functional theory-DFT, UV/Vis spectrophotometry, AFM, Raman spectroscopy, and SERS). The resulting gold(III) complexes/AuNPs conjugates turned out to be pretty stable. Interestingly, they exhibited a dramatically increased resonance intensity in the Raman spectra induced by AuNPs. For testing the use of the functionalized AuNPs for biosensing, their distribution in the nuclear, cytosolic, and membrane cell fractions obtained from human lymphocytes was investigated by AFM and SERS. The conjugates were detected in the membrane and nuclear cell fractions but not in the cytosol. The AFM method confirmed that conjugates induced changes in the morphology and nanostructure of the membrane and nuclear fractions. The obtained results point out that the conjugates formed between AuNPs and gold(III) complexes may be used as a tool for tracking metallodrug distribution in the different cell fractions.

Interaction of Au(iii) and Pt(ii) complexes with Na/K-ATPase: experimental and theoretical study of reaction stoichiometry and binding sites.

The present paper deals with investigation of the interaction between selected simple structure Au(iii) ([AuCl4]-, [AuCl2(dmso)2]+, [AuCl2(bipy)]+) and Pt(ii) ([PtCl2(dmso)2]) complexes with Na/K-ATPase as the target enzyme, using an experimental and theoretical approach. Reaction stoichiometries and binding constants for these enzyme/complex systems were determined, while kinetic measurements were used in order to reveal the type of inhibition. Based on the results obtained by quantum mechanical calculations (electrostatic surface potential (ESP), volume and surface of the complexes) the nature of the investigated complexes was characterized. By using the solvent accessible surface area (SASA) applied on specific inhibitory sites (ion channel and intracellular domains) the nature of these sites was described. Docking studies were used to determine the theoretical probability of the non-covalent metal binding site positions. Inhibition studies implied that all the investigated complexes decreased the activity of the enzyme while the kinetic analysis indicated an uncompetitive mode of inhibition for the selected complexes. Docking results suggested that the main inhibitory site of all these complexes is located in the ion translocation pathway on the extracellular side in the E2P enzyme conformation, similar to the case of cardiac glycosides, specific Na/K-ATPase inhibitors. Also, based on our knowledge, the hydrolyzed forms of [AuCl4]- and [PtCl2(dmso)2] complexes were investigated for the first time by theoretical calculations in this paper. Thereby, a new inhibitory site situated between the M2 and M4 helices was revealed. Binding in this site induces conformational changes in the enzyme domains and perturbs the E1-E2P conformational equilibrium, causing enzyme inhibition.