Structural and functional studies of heavy metal ATPases

Publikation: Bog/antologi/afhandling/rapportPh.d.-afhandlingForskning

Dokumenter

  • PhD_thesis

    Accepteret manuskript, 49,5 MB, PDF-dokument

  • Oleg Sitsel
Copper and zinc are trace elements that are crucial for the well-being of all cells and are an indispensable part of many proteins. At the same time, the intracellular levels of these metals require careful regulation, as an excess or deficiency may be lethal. P1B-ATPases are key players in Cu+ and Zn2+ homeostasis that belong to the superfamily of P-type ATPases, transmembrane proteins which are present in virtually all lifeforms, with functions ranging from membrane potential generation to muscle relaxation. The goal of this thesis is to improve our understanding of P1B-ATPases by focusing on the bacterial proteins LpCopA and SsZntA, which represent Cu+- and Zn2+-ATPases, respectively. The thesis first compares the recent pioneering P1B-ATPase structure of LpCopA to that of the well-described Ca2+-ATPase SERCA, showing how Cu+-ATPases have managed to adapt the general P-type ATPase topology to handle heavy metal ions. LpCopA is then compared to its two human homologues ATP7A and ATP7B, which cause the severe Menkes and Wilson diseases when malfunctioning. The differences between the three proteins are described and disease-causing mutations in the human proteins are analyzed. The crystal structure of LpCopA in a new conformational state is then presented and studied using a variety of methods, showing that Cu+-ATPases use an ion release pathway unique for the P-type ATPase superfamily. The next section introduces the two pioneering crystal structures of a Zn2+-ATPase, SsZntA. Investigation of SsZntA using a broad range of techniques reveals an array of unique Zn2+-ATPase features relating to ion uptake, binding, discharge and countertransport. These results are then used to comprehensively compare Zn2+-ATPases with their Cu+-transporting counterparts, showing how the two have managed to adapt to transport of distinct ion types while still maintaining all the major features of the P1B-ATPase subclass.
OriginalsprogEngelsk
Antal sider236
Rekvirerende organGraduate School of Science and Technology
StatusUdgivet - 28 aug. 2015

Note vedr. afhandling

During his PhD studies, Oleg Sitsel has worked on proteins called P1B-ATPases which transport heavy metals across cell membranes, the barriers which separate cells and intracellular compartments from their immediate environment. P1B-ATPases are important for correct regulation of cellular levels of metals such as copper and zinc, which are crucial for the well-being of all cells and are an indispensable part of many proteins. At the same time, an excess or deficiency of these elements can be lethal, as can be seen in the case of the human Menkes and Wilson diseases caused by malfunction of copper transporting P1B-ATPases.

Oleg Sitsel has studied copper and zinc transporting P1B-ATPases using a broad array of biochemical and biophysical techniques such as X-ray crystallography in order to obtain insight on how these proteins work at an atomic level. The research he conducted resulted in the discovery of an ion release pathway unique to copper transporting P1B-ATPases, as well as the first ever structures of a zinc transporting P1B-ATPase. These major breakthroughs in the field give hope for using these proteins in a range of new applications, from developing new antibiotics to creating heavy metal extracting plants.

The PhD degree was completed at the Department of Molecular Biology and Genetics, Science and Technology, Aarhus University

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