On 13th of July, 2022, from 11:00 to 12:20 CET, the fifth meeting of the calixDNA research team was held, in presence of A. Višnjevac, I. Piantanida, O. Reinaud, B. Kojić-Prodić, K. Gruber, I. Nikšić-Franjić, D. Pavlović Saftić, M. Wallet and M. Legras. M. Wallet and M. Legras presented the results obtained during their internship at the calixDNA project, which started on April 23rd, 2022, and will continue until the end of July 2022, which was followed by the vivid discussion and many questions to the presenters. Research stay of I.Nikšić-Franjić at the research group of prof. K. Gruber was confirmed for the late September/early October, during which she will set up the first round of crystallization experiments of the genZ SBB4 – DNA oligomere complexes. Research visits of A. Višnjevac and I. Nikšić Franjić to the Paris lab, where they will continue the synthesis of the gram quantities of the most promissing SBB derivatives, was also mentioned and confirmed.
Board of Directors of the Croatian Science Foundation, at its 27th session held on June 17th, 2022, based on the evaluations and recommendations of the evaluators, made a decision (Class: 120-02/22-02/13, Registration number: 63-02/ 01-22-33) on the acceptance of the 1st periodic report of calixDNA project (Feb. 1st, 2021 – Jan. 31st, 2022). Final grade B (Good progress) has been assigned to the report with the remark that “the project has achieved most of the goals for the given period, and the reasons for not achieving some goals are justified”. Consequently, the board has decided to continue financing the project.
calixDNA project was presented to the widest audience in Croatia by the well known science&technology dedicated TV show “Prometej” which was broadcasted on 4th od June, 2022 at 2:30 PM at HRT1. The cameras of the HRT entered the Laboratory of Chemical and Biological Crystallograhy and Laboratory for biomolecular interactions and spectroscopy at the Ruđer Bošković Institute to get an insight into the research process within the calixDNA project. The project was presented by its head, dr. sc. Aleksandar Višnjevac, and particular research activities by dr. sc. Ivana Nikšić-Franjić, Malorie Wallet and Marine Legrass, team members.
The abstract entitled Impact of positive charge and ring-size on interactions of calixarenes with DNA, RNA and nucleotides by Ivana Nikšić-Franjić, Aleksandar Višnjevac and Ivo Piantanida has been accepted for an oral presentation at the EuChemS2022 Chemistry Congress. Presenting author is dr. Ivana Nikšić-Franjić, postdoc fellow at the calixDNA.
Calixarenes were traditionally studied as supramolecular receptors for various low molecular mass cationic or anionic species, but only marginally investigated for their biological effects or applications . However, systematic study revealed that cationic calixarenes can be applied as innovative DNA-transfection agents . We have developed three generations of calixarene-based (funnel), as well as two generations of resorcinarene-based (bowl) supramolecular systems featuring two, three or four methylimidazole-containing coordination arms grafted at the large rim, suitable to bind to variety of nucleid acid sequences [3,4].
In this work we focused on calixarenes with short, triazole-attached positively charged substituents (2 and 4) and their neutral analogues (1 and 3, Fig. 1.). Their ability to recognize specific sequences of various DNA chains was studied by thermal denaturation experiments, fluorimetric titrations, circular dichroism experiments and molecular dynamics simulations. Comparison of neutral and cationic calixarene and calixarene derivatives revealed that only cationic analogues non-covalently bind to ds-DNA and ds-RNA, by insertion into DNA minor groove or RNA major groove. Also, cationic analogues revealed strong and highly selective charge-dependent stabilization of AT-DNA against thermal denaturation, both neutral and cationic calixarenes bind nucleoside monophosphates with similar efficiency, by forming tweezer-like supramolecular complexes, with nucleobases inserted between aromatic pendant arms grafted to calixarene rims. Such nucleotide-calixarene complexes were monitored by emission change of calixarene as a function of nucleobase insertion, at variance to DNA/RNA complexes in which calixarene is inserted into polynucleotide groove, which do not change calixarene emission – stressing importance of the ligand insertion within calix-basket for the fluorimetric sensing.
calixDNA team is joined, from April 23rd, 2022, by Marine Legras and Malorie Wallet, students of the second year of chemistry at the ENSICAEN (Ecole Nationale Superieure de Caen). During their internship at the Ruđer Bošković Institute, they will be working on the recognition and binding studies of the gen1 supramolecular biomimetic binders, as well as on the series of crystallization experiments pertaining to the structural studies of the genZ SBB- mononulceotide tweezer like complexes, and metal complexes of some of the genZ SBBs. Both Malorie and Marine will stay with us for a period of three months.
Our article entitled Impact of positive charge and ring-size on interactions of calixarenes with DNA, RNA and nucleotides by Ivona Krošl, Ena Otković, Ivana Nikšić-Franjić, Benoit Colasson, Olivia Reinaud, Aleksandar Višnjevac and Ivo Piantanida, has been accepted for publication in New Journal of Chemistry. The publication summarizes part of the work accomplished during the first product period, on the recognition and binding study of the selected SBVs towards variety of polynucleotide chains and some mononucleotides.
Comparison of various calixarene and calixarene derivatives revealed that only analogues bearing permanent positive charge non-covalently bind to ds-DNA and ds-RNA, by insertion into DNA minor groove or RNA major groove. Also, these cationic analogues revealed strong and highly selective charge-dependent stabilization of AT-DNA against thermal denaturation, calixarene trication being for an order of magnitude more efficient than its calixarene dicationic analogue. At variance to DNA/RNA selectivity for only cationic calixarenes, both, neutral and cationic calixarenes bind nucleoside monophosphates with similar efficiency, by forming tweezer-like supramolecular complexes, with nucleobases inserted between aromatic pendant arms grafted to calixarene rims. Such nucleotide-calixarene complexes were monitored by emission change of calixarene as a function of nucleobase insertion, at variance to DNA/RNA complexes in which calixarene is inserted into polynucleotide groove, which does not change calixarene emission – stressing importance of the ligand insertion within calix-basket for the fluorimetric sensing.
We are, again, looking for motivated students interested in joining our international calixDNA team in frame of their BSc/MSc theses. An ideal candidate would be a student of chemistry or biochemistry, with a strong interest in structural features of the biologically relevant molecules, as well as biological macromolecules. We offer a position in the field of the recognition and binding studies of our supramolecular biomimetic binders (SBBs) to the oligonucleotide chains. Experimental work will be organized at the Ruđer Bošković Institute in Zagreb, Bijenička c. 54.
The project is ongoing, hence your experimental work can start immediately. Please visit calixdna.org for all the details you may be interested in. Should you have any further inquiries, there is a contact form on the website, through which you can reach out. We would be delighted to welcome you on board calixDNA project.
On Sep. 22nd, 2021, the first MSc student at the calixDNA project, Ms Ena Otković, from the University of Zagreb, Faculty of Sciences, successfully defended her thesis of the title STUDY OF CATIONIC CALIXARENES INTERACTIONS WITH NUCLEOTIDES, DNA AND RNA, accomplished under the supervision of dr. sc. Ivo Piantanida. The thesis was defended before the evaluation committee, president of which was prof. Snežana Miljanić, members were profs. Željka Soldin and Marko Močibob, while prof. Adriana Kenđel was appointed first replacement. Thesis is written in Croatian, with an abstract available in English, and is deposited in Central Chemical Library, Faculty of Science, University of Zagreb, Horvatovac 102a, Zagreb, Croatia and in Repository of the Faculty of Science, University of Zagreb. Full text in Croatian is also available on the calixDNA website.
The calixDNA project team had today its 4th online meeting from 2:00 PM to 3:30 PM, in presence of A. Višnjevac, O. Reinaud, B. Colasson, I. Piantanida, I. Krošl, I. Nikšić-Franjić and C. Thouzé. The central point on the agenda was the final internship report of Corentin Thouzé, who will terminate his internship at the calixDNA project on Aug. 13th, 2021, as planned. Corentin has wrapped up and presented, in a form of a 35 min long PPT presentation, the results obtained so far on the research of the binding of two calixarene derivatives and their corresponding Cu(II) complexes (supramolecular biomimetic binders, SBBs) to various mono- and polynucleotides. The research involved UV/Vis, fluorometric and CD spectroscopies, as well as thermal denaturation monitoring, which, combined, provided a detailed insight into the chemical behaviour of the studied SBBs in presence of variety of polynucleotide chains, as well as in presence of their building blocks – mononucleotides. The results were put in context of the previously obtained ones, on the series of four closely related SBBs, which are the basis of the MSc thesis of Ena Otković, and of the first calixDNA original scientific paper to be submitted shortly to the special issue of Molecules – an open access journal. The presentation incited a vivid scientific discussion, which, in many aspects, contributed to better understanding of the presented results. One of the conclusions was that Corentin shall continue the research on Ena’s SBBs, by studying binding ability of their copper(II) complexes to polynucleotide chains.
We have briefly discussed the paper in preparation (vide supra) and current project management issues. French team members will, over the two weeks to come, review the manuscript and send their comments and suggestions, so that the paper will be ready for submission in August 2021. A. Višnjevac will, if the circumstances remain favorable to international visit exchange, visit the Lab at the University of Paris, during November/December 2021, in frame of the joint team efforts to speed up the synthesis of the zero generation SBBs, needed for the envisaged recognition, binding and structural studies during the second project year.