Speaker: Christine Payne
Department: School of Chemistry and Biochemistry, Georgia Tech
Subject: Material-cell interactions
Location: TU Delft
Date: 15 October 2015
Author: Gabriele Kockelkoren
The goal of research in the Payne Lab at Georgia Tech is to understand the underlying molecular mechanisms by which cells interact with materials. This talk was divided into the two main research topics of her lab: nanoparticles interactions and conducting polymers. In her research the use of fluorescence microscopy for cellular interactions to get the spatial form of the cell is essential. Professor Payne makes also use of spectroscopy at the molecular level and calorimetry.
Nanoparticles can be used for cellular applications as drug delivery of imagining and sensing. Her lab focuses more on these last two applications. As her lab is interested in the interactions of nanoparticles with cells, cells are cultured in a aqueous solution of salts, amino acids and glucose. In the medium there is also the serum FBS. How surface of nanoparticles affects binding of FBS. They have one cationic nanoparticles and one anionic nanoparticles. They use a nanoparticle washing procedure. At some wash you do not see the protein anymore. They found albumin on the surface. It didn’t matter what the surface charge was, both experiments gave the same result for albumin presence.
These images are made using fluorescence microscopy. The left part of the image shows cationic nanoparticles and the right part anionic nanoparticles. Some cells are placed in only MEM, others are placed in a MEM medium including a 10% FBS solution. Cationic NPs are amine-modified and anionic NPs are carboxylate-modified. Source: Fleischer CC, Payne CK. Nanoparticle surface charge mediates the cellular receptors used by protein-nanoparticle complexes. J Phys Chem B. 2012;116:8901–8907.
The Pain-lab continued its researched the effects of FBS on the binding properties of NPs at the cell. For this they have used ionic and cationic NPs. To look at the interactions at the cellular level epi-fluorescent microscopy was used. Cellular binding is a function of the initial NP charge. If nanoparticles are initially cationic, the no particles can be observed. If a certain percentage of FBS is present, then NPs are to be seen. The experiments showed that for anionic NPs, FBS inhibits the binding of nanoparticles. For this experiment special NP-washing procedures have been used. To check their results, they compared these to experiments with BSA. The identical results were found. It can be seen that NPs are competing with the free proteins for the binding site. Complexes that are formed from anionic NPs use native protein receptors. Furthermore it is shown that increasing the FBS concentration, decreases the binding affinity. In summary, this experiment showed that cationic NPs with BSA use a different cellular receptor than BSA, while anionic use the same receptor. The lab had already hypothesized which receptor this could be. To test whether indeed anionic NPs bind to this specific receptor, a competition essay between polyinosinic acid which binds the receptor and NPs was done. Using CD spectrometry it was checked whether there where structural changes in the protein when it was bound to the cell surface. For anionic NPs we see no change, however for ionic NPs a clear shift can be seen in spectrometry. These experiments led to the conclusion that the protein corona structure determines which cell surface receptors can be used for the binding of NPs.
In the second part of the seminar conducting polymer-cell interactions were highlighted. Mrs Payne and her colleagues want to synthesize PEDOT:PSS in cells, this is the biomolecular synthesis of conducting polymers. PEDOT:PSS are highly conductive polymers made out of the PEDOT monomer and the PSS monomer. Peroxisomes control intracellular reactions and are organelles where fatty acids are degraded in the cell. Most importantly, PEDOT:PSS is synthesized here. Peroxisomes contain a catalyst, called catalase. By depositing monomers in these peroxisomes, it has been shown that PEDOT:PSS is formed. By firstly boiling catalase (so that it loses its enzymatic activities), the experiment has been repeated several times. Even though the enzyme was denatured, still PEDOT:PSS was produced. This indicates that enzyme activity is not required. This experiment has been done with other proteins as well, like transferrin. Transferrin is associated with transport of iron in the blood. The results were exactly the same, PEDOT:PSS was produced. This shows that enzymatic activity is not needed for PEDOT:PSS formation, however an iron containing protein in necessary for the reaction to work. In contract to nowadays techniques to form PEDOT:PSS, this method allows formation in a single step.
I liked this seminar very much. All kinds of research techniques were discussed, like spectrometry, quantum dots and fluorescence microscopy. It is very nice to recognize those techniques and know their basics work.