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315 464-8703

Stephan Wilkens, PhD

4237 Weiskotten Hall
766 Irving Avenue
Syracuse, NY 13210
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Associate Professor of Biochemistry and Molecular Biology




Biochemistry and Molecular Biology
Biomedical Sciences Program
Cancer Research Program


Structure and Mechanism of Membrane Bound Transport Proteins


Postdoctoral Fellow: University of Oregon, Institute of Molecular Biology
PhD: Freie University of Berlin, 1995


figure 1Our main focus is on the structure and mechanism of the eukaryotic proton pumping vacuolar ATPase (V-ATPase; V1Vo-ATPase). V-ATPase is a large, multisubunit membrane protein complex found on the endomembrane system of all eukaryotic cells, where it functions to acidify intracellular organelles, including endosomes, lysosomes, the Golgi apparatus and synaptic vesicles. An acidic pH in the lumen of these compartments is essential for basic cellular functions including endocytosis, protein degradation, protein trafficking, and neurotransmitter loading. When found on the plasma membrane of "professional" acid secreting cells such as bone osteoclasts and renal intercalated cells, V-ATPase pumps protons to the outside of the cell, a process required for bone remodeling and urine acidification. Due to its involvement in numerous basic cellular functions, a malfunctioning V-ATPase can lead to widespread human diseases including osteoporosis, neurodegeneration, diabetes, cancer, and AIDS.

figure 2V-ATPase is composed of two rotary motor subcomplexes, a membrane extrinsic ATP hydrolyzing V1-ATPase, and a membrane embedded Vo proton turbine. ATP hydrolysis on V1 is coupled to proton transport across Vo by a "central stalk" that rotates up to 100 1/s to pick up protons from the cytosol and deliver them to the organelle lumen or outside of the cell. As a major consumer of cellular energy, V-ATPase function is tightly regulated by a mechanism referred to as "reversible disassembly". When the enzyme's proton pumping is not needed, V1 dissociates from the membrane bound Vo, and the activity of both subcomplexes is "silenced" so that V1 is no longer capable of hydrolyzing MgATP, and Vo becomes impermeable to protons. Reversible disassembly is unique to eukaryotic V-ATPase and the process offers a way to modulate the activity of a disease causing V-ATPase for therapeutic purposes.

figure 3We work with the V-ATPases from the model organism Saccharomyces cerevisiae (baker's yeast) as well as human. We are using the tools of biochemistry, molecular & cell biology, biophysics and structural biology (including NMR spectroscopy, X-ray crystallography and cryo electron microscopy (cryoEM)) to obtain structural and mechanistic information. Our long term goal is to elucidate the mechanism of V-ATPase's unique mode of regulation and find ways for therapeutic targeting of the enzyme in the disease state.

Recent Publications

Sharma, S., Oot, R.A., Khan, Md M. and Wilkens, S. (2019) Functional reconstitution of vacuolar H+-ATPase from Vo proton channel and mutant V1-ATPase provides insight into the mechanism of regulation by reversible disassembly. J. Biol. Chem., in press.

Sharma, S., Oot, R.A. and Wilkens, S. (2018) MgATP hydrolysis destabilizes the interaction between subunit H and yeast V1-ATPase, highlighting H's role in V-ATPase regulation by reversible disassembly. J. Biol. Chem. 293, 10718-10730.

Roh, S.-H., Stam, N.J., Hryc, C., Couoh-Crdel, S., Pintillie, G, Chiu, W. and Wilkens, S. (2018) The 3.5-Å CryoEM Structure of Nanodisc-Reconstituted Yeast Vacuolar ATPase Vo Proton Channel. Mol. Cell 69, 993-1004.

Oot, R.A., Couoh-Cardel, S., Sharma, S., Stam, N.J. and Wilkens, S. (2017) Breaking up and Making up: The Secret Life of the Vacuolar H+-ATPase. Protein Science 26, 896-909.

Sharma, S. and Wilkens, S. (2017) Biolayer Interferometry of Lipid Nanodisc-Reconstituted Yeast Vacuolar H+-ATPase. Protein Science 26, 1070-1079.

Stam, N.J. and Wilkens, S. (2017) Structure of Nanodisc Reconstituted Vacuolar ATPase Proton Channel: Definition of the Interaction of Rotor and Stator and Implications for Enzyme Regulation by Reversible Dissociation. J. Biol. Chem. 292, 1749-61.

Oot, R. A., Kane, P.M., Berry, E.A. and Wilkens, S. (2016) Crystal Structure of Yeast V1-ATPase in the Autoinhibited State. EMBO J. 35,1694-706.

Couoh-Cardel, S., Hsueh, Y.-C., Wilkens, S. and Movileanu, L. (2016) Yeast V-ATPase Proteolipid Ring Acts as a Large-Conductance Transmembrane Protein Pore. Sci. Rep. 6, 24774.


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