- Compatible solute uptake
- Mechanosensitive channels
- Diffusional barriers
Compatible solute uptake (Prof. Dr. B. Poolman)
ABC (ATP binding cassette) transporters catalyze the transport of a wide variety of molecules across lipid bilayers into or out of cells and organelles. They form one of the largest known protein families and are found in organisms from all kingdoms of life. The basic architecture of ABC transporters1,2 is shown in the figure. One of the ABC transporters that is studied in detail is the osmoregulatory ABC transporter OpuA (class D). Osmotic control of OpuA involves gating by intracellular ionic strength and is mediated by lipid-protein interactions.3 The tandem CBS domain4, which is linked to the nucleotide binding protein, plays a pivotal role in the regulation. A structural genomics approach is used to determine to the structures of proteins belonging to other classes of ABC transporters.
Heide van der T., and Poolman, B. (2002) ABC transporters: one, two or four extracytoplasmic substrate-binding sites? EMBO Rep., 3, 938-943.
Biemans-Oldehinkel, H., Doeven, M.K., and Poolman, B. (2006) ABC transporter architecture and regulatory roles of accessory domains. FEBS Lett. , 580 , 1023-1035 .
Heide, T. van der, Stuart, M.C.A., and Poolman, B. (2001) On the osmotic signal and osmosensing mechanism of an ABC transport system for glycine betaine.
EMBO Journal, 20, 7022-7032.
Biemans-Oldehinkel, H., Mahmood, N.A.B.N., and Poolman, B. (2006) A sensor for intracellular ionic strength. Proc. Natl. Acad. Sci. USA , 103, 10624-10629.
Mechanosensitive channels (Prof. Dr. B. Poolman, Dr. A. Kocer)
Whereas osmoregulatory ABC transporters protect cells against hyperosmotic stress, mechanosensitive channels like MscL offer protection against osmotic downshifts. MscL is gated by changes in the tension in the membrane. We have shown that, in the open state MscL, forms pores with a diameter of 3 nm1, which allows passage of small proteins. Synthetic biology approaches are used to gate MscL photochemically by incorporating switches in the surrounding lipids2 or at constriction site of the pore (‘charging’ the pore3,4).
- van den Bogaart, G., Krasnikov, V., and Poolman, B. (2006) Dual-color fluorescence burst analysis to probe protein diffusion through the mechanosensitive channel MscL. Biophys. J ., 92, 1233-1240.
- Folgering, J.H.A., Kuiper, J.M., de Vries, A.H., Engberts, J.B.F.N., and Poolman, B (2004) Lipid-mediated light-activation of a mechanosensitive channel of large conductance. Langmuir 20, 6985-6987.
- Kocer, A., Walko, M., Meijberg, W., and Feringa, B.L. (2005) A light-actuated nanovalve derived from a channel protein. Science. 309, 755-758.
Kocer, A., Walko, M., and Feringa, B.L. (2007) Synthesis and utilization of reversible and irreversible light-activated nanovalves derived from the channel protein MscL. Nature Protoc., 2, 1426-1437.
Diffusional barriers (Prof. Dr. B. Poolman)
We investigate the mobility of proteins in the bacterial cytoplasm and cytoplasmic membrane as well as membrane model systems, using optical microscopy-based techniques such as fluorescence correlation spectroscopy (FCS) and pulsed-FRAP (panel a). How does crowding affect mobility of proteins and lipids? Are diffusional barriers formed when the crowding gets too high? We have proposed that under conditions of severe osmotic stress (panel b, lower figure), the cell membrane contacts the nucleoid, which creates pools of proteins and hinders free diffusion through the cell.
Kahya, N., Scherfeld, D., Bacia, K., Poolman, B., and Schwille, P. (2003)
Probing lipid mobility of raft-exhibiting model membranes by Fluorescence Correlation Spectroscopy.
J. Biol. Chem., 278, 28109-28115.
van den Bogaart, G., Hermans, N., Krasnikov, V., and Poolman, B. (2007) Protein mobility and diffusive barriers in Escherichia coli: consequences of osmotic stress. Molec. Microbiol., 64, 858-871.
|Last modified:||03 October 2012 12.04 p.m.|