Amongst bacteria, living in complex and varying communities and surroundings, rivalry for crucial resources exists. Due to the constant pressure, strategies to allow competition and communication have been developed within bacterial communities. In this respect, Contact Dependent Growth Inhibition (CDI) is often of relevance. The aim was to elucidate a newly found pathway of transport of the CDI toxins across the inner bacterial membrane via Sec translocon. Since all proteins are transported via Sec translocon in an unfolded state, and there is no obvious mechanism which would drive or facilitate this transport in the retrograde fashion, the hypothesis that the effector domain of CDI toxins evolved an anisotropic energy landscape of mechanical unfolding was constructed. That would allow the toxin to be mechanically more labile in the direction of translocation and mechanically stable in the orthogonal directions. This anisotropy would permit efficient translocation and overall thermodynamic stability at the same time. The unfolding landscape was assessed by a molecular dynamics simulation combined with umbrella sampling. Developed methodology, complemented with a rational design of "circular permutants", helped to estimate the forces required to unfold the toxins under different geometries.
Anotace v angličtině
Amongst bacteria, living in complex and varying communities and surroundings, rivalry for crucial resources exists. Due to the constant pressure, strategies to allow competition and communication have been developed within bacterial communities. In this respect, Contact Dependent Growth Inhibition (CDI) is often of relevance. The aim was to elucidate a newly found pathway of transport of the CDI toxins across the inner bacterial membrane via Sec translocon. Since all proteins are transported via Sec translocon in an unfolded state, and there is no obvious mechanism which would drive or facilitate this transport in the retrograde fashion, the hypothesis that the effector domain of CDI toxins evolved an anisotropic energy landscape of mechanical unfolding was constructed. That would allow the toxin to be mechanically more labile in the direction of translocation and mechanically stable in the orthogonal directions. This anisotropy would permit efficient translocation and overall thermodynamic stability at the same time. The unfolding landscape was assessed by a molecular dynamics simulation combined with umbrella sampling. Developed methodology, complemented with a rational design of "circular permutants", helped to estimate the forces required to unfold the toxins under different geometries.
Amongst bacteria, living in complex and varying communities and surroundings, rivalry for crucial resources exists. Due to the constant pressure, strategies to allow competition and communication have been developed within bacterial communities. In this respect, Contact Dependent Growth Inhibition (CDI) is often of relevance. The aim was to elucidate a newly found pathway of transport of the CDI toxins across the inner bacterial membrane via Sec translocon. Since all proteins are transported via Sec translocon in an unfolded state, and there is no obvious mechanism which would drive or facilitate this transport in the retrograde fashion, the hypothesis that the effector domain of CDI toxins evolved an anisotropic energy landscape of mechanical unfolding was constructed. That would allow the toxin to be mechanically more labile in the direction of translocation and mechanically stable in the orthogonal directions. This anisotropy would permit efficient translocation and overall thermodynamic stability at the same time. The unfolding landscape was assessed by a molecular dynamics simulation combined with umbrella sampling. Developed methodology, complemented with a rational design of "circular permutants", helped to estimate the forces required to unfold the toxins under different geometries.
Anotace v angličtině
Amongst bacteria, living in complex and varying communities and surroundings, rivalry for crucial resources exists. Due to the constant pressure, strategies to allow competition and communication have been developed within bacterial communities. In this respect, Contact Dependent Growth Inhibition (CDI) is often of relevance. The aim was to elucidate a newly found pathway of transport of the CDI toxins across the inner bacterial membrane via Sec translocon. Since all proteins are transported via Sec translocon in an unfolded state, and there is no obvious mechanism which would drive or facilitate this transport in the retrograde fashion, the hypothesis that the effector domain of CDI toxins evolved an anisotropic energy landscape of mechanical unfolding was constructed. That would allow the toxin to be mechanically more labile in the direction of translocation and mechanically stable in the orthogonal directions. This anisotropy would permit efficient translocation and overall thermodynamic stability at the same time. The unfolding landscape was assessed by a molecular dynamics simulation combined with umbrella sampling. Developed methodology, complemented with a rational design of "circular permutants", helped to estimate the forces required to unfold the toxins under different geometries.
Dr Štěrba welcomed the student and commission members; Prof Vácha was not present. The student presented the theoretical background of her work on the transport system the studied proteins and the used methods, followed by the experimental part and the results. The supervisor, Dr Fessl, and the opponent, Dr. Franta, presented their reviews. The student answered all the questions of the opponent and also questions from the commission members regarding future experiments, additional possible measurements, the requirement of energy for unfolding and refolding and the conditions for refolding of the transported proteins.
Votes: 4x 1
Final grade: 1
Points: 145
The commission recommends this thesis to be awarded by the head of the department.