Molecular modelling and experimental studies of the interactions between biomolecules and nanostructured inorganic materials

In this study the interactions of biomacromolecules with different inorganic nanostructured materials were investigated using experimental and theoretical approaches. A deeper understanding on the fundamental steps involved for such interactions is very important for the fields of nanotechnology, medicine and biotechnology. In nanotechnology biomolecules like DNA have often been considered as templates for the scaffolding of nanoparticles. TEM observations of DNA molecules with Au55 cluster indicated the formation of equidistant gold nanowires. Interestingly the nanowires were composed of such dimensions that indicated that the Au55 cluster initially used was reduced to the size of a cluster with 13 gold atoms. In order to understand the formation of gold nanowires, various molecular modelling techniques were employed. Based on molecular mechanics simulations, it was found that Au55 cluster energetically prefers to interact with the major groove of B-DNA whereas Au13 prefers to associate with the major grooves of A-DNA. It is not clear how the reduction of Au55 to Au13 occurs however it is assumed that it is influenced by the dramatic alterations occurring in the groove geometry during B-DNA to A-DNA transition. The formation of gold nanowires is determined by the equidistant decoration of Au13 clusters around the major grooves of A-DNA. Molecular dynamics simulations indicated that the aggregation of such DNA-gold complexes leads to the formation of nanowires which are separated from each other almost 0.5 nm. The ability of Au55 to interact with the major grooves of B-DNA led to further investigations on the potential of this gold cluster as an anticancer drug. Gold-based compounds have not been studied as extensively as other metal-based drugs like platinum derivatives which have been employed as chemotherapeutic agents for many years in medical oncology. A variety of human cancer and healthy cell lines have been employed for investigating sensitivity towards Au55 and cisplatin. In vitro cytotoxicity assays performed by a colourimetric-based method (MTT) have indicated that generally Au55 is more toxic and efficient than cisplatin. Especially the cisplatin resistant metastatic melanoma cell lines exhibited the highest sensitivity towards Au55. Apoptotic or necrotic death influenced by Au55 was investigated by performing a fluorometric assay which measures the induction of the apoptotic proteins caspases –3 and –7. The effect of Au55 was different among the five cell lines investigated. In the case of the osteosarcoma cell line U-2OS induction of caspases –3 and –7 in the presence either of Au55 or cisplatin was observed indicating apoptosis being triggered. In the other four different cell lines the induction of caspases either in the presence of Au55 or cisplatin was similar to the controls. This indicates that these cells could be dying either by necrosis, or apoptosis is induced through a caspase-independent pathway. Further information on the cellular localisation and possible target has been investigated by measuring the distribution of radioactive gold cluster in various cellular fractions of the BLM cell line. It has been found that slightly higher amount of Au55 was located in the chromosomal DNA rather than in the cytoplasmic fraction. The Au55 was strongly bound to the chromosomal DNA with a very small fraction of it being present in the nuclear proteins. This has indicated that DNA is one of the main cellular targets however it is not certain whether such an interaction is directly related to the cytotoxic effect of Au55. The gold cluster could possibly damage the chromosomal DNA in the presence of H2O2 as indicated by the conversion of supercoiled plasmid DNA into open-circular form by agarose gel electrophoresis. Molecular docking simulations have shown the energetic preference of this agent for AATT sequences which is different to the GG and AG preference of cisplatin. Finally protein-gold cluster interactions could also play an important role on the cytotoxic effect of Au55. In vitro cytotoxicity assays in the presence of albumin have shown an increase in the IC50 values indicating a sequestering effect of albumin protein. Molecular docking simulations suggest the interaction of Au55 with specific cystein residues of human albumin. Nanostructured inorganic materials have been considered very important in the field of biotechnology with major applications aiming towards the immobilisation or purification of biomolecules such as proteins and amino acids. In this study the interactions of three different amino acids with a zeolite structure have been investigated using molecular modelling. MD simulations have been performed in the presence of water molecules with Phe, Lys and Glu being placed around the structure of zeolite beta. The simulations have indicated that amino acids energetically prefer to interact with the surface of the zeolite-beta rather being placed inside the pores or in the bulk. MD simulations clearly indicate the preference of Phe to adsorb through the formation of hydrogen bonds with the O atoms of the zeolite structure. The amino acids Lys and Glu have indicated a lower preference as the formation of fewer hydrogen bonds with the zeolite surface has been exhibited. This is in aggreement with experimental studies reported in the literature. Nevertheless MD simulations have not been able to distinguish any differences on the adsorption behavior of Glu and Lys.


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