The cell membrane is a biological membrane that separates the interior of all cells from the outside environment which protects the cell from its environment consisting of a lipid bilayer with embedded proteins. The cell membrane controls the movement of substances in and out of cells and organelles. In this way, it is selectively permeable to ions and organic molecules. In addition, cell membranes are involved in a variety of cellular processes such as cell adhesion, ion conductivity and cell signalling and serve as the attachment surface for several extracellular structures, including the cell wall, the carbohydrate layer called the glycocalyx, and the intracellular network of protein fibers called the cytoskeleton.
Membranes protect all of our cells and the organelles inside them, including the mitochondria – the powerhouse of the cell. These membranes are studded with biological machinery made of proteins that enable molecular cargo to pass in and out.
Studying these membrane-embedded machines in their native state is crucial to understanding mechanisms of disease and providing new goals for treatments. However, current methods for studying them involve removing them from the membrane, which can compromise the integrity of membrane proteins and alter their structure and functional properties.
Recently, researchers have developed a new technique to analyse cell membrane proteins in situ which could revolutionise the way in which we study diseases, such as cancer, metabolic and heart diseases. In this work, they ejected intact assemblies from membranes, without chemical disruption, and used mass spectrometry to define their composition. This research will enable the development of mass spectrometry in biology to be taken to a new level, enabling new discoveries that would not have been possible before.
The technique could dramatically affect our understanding of both how cell membrane complexes work, and in the process, our approach to healthcare research.
From Escherichia coli outer membranes, scientists identified a chaperone-porin association and lipid interactions in the β-barrel assembly machinery. They observed efflux pumps bridging inner and outer membranes, and from inner membranes they identified a pentameric pore of TonB, as well as the protein-conducting channel SecYEG in association with F1FO adenosine triphosphate (ATP) synthase. Intact mitochondrial membranes from Bos taurus yielded respiratory complexes and fatty acid–bound dimers of the ADP (adenosine diphosphate)/ATP translocase (ANT-1). These results highlight the importance of native membrane environments for retaining small-molecule binding, subunit interactions, and associated chaperones of the membrane proteome.
The technique involves vibrating the sample at ultrasonic frequencies so that the cell begins to fall apart. Electrical currents then applied an electric field to eject the protein machines out of the membrane and directly into a mass spectrometer – an instrument that can detect a molecule’s chemical ‘signature’, based on its mass.
Not only did the membrane protein machines survive the ejection; the analysis also revealed how they communicate with each other, are guided to their final location and transport their molecular cargo into the cell. With the development of this method, the application of mass spectrometry in biology will be taken to a new level, using it to make discoveries that would not have been possible before.
“A longstanding question on the structure of one membrane machine from mitochondria has now been solved using this technique.” said Dr Sarah Rouse, from the Department of Life Sciences at Imperial. The results are particularly exciting for mitochondrial membranes—we managed to catch a translocator in action—passing metabolites. Because mitochondrial therapeutics target a wide range of debilitating diseases, we now have a new way of assessing their effects.
Title: Protein assemblies ejected directly from native membranes yield complexes for mass spectrometry
Authors: Dror S. Chorev, Lindsay A. Baker, Di Wu, Victoria Beilsten-Edmands, Sarah L. Rouse, Tzviya Zeev-Ben-Mordehai, Chimari Jiko, Firdaus Samsudin, Christoph Gerle, Syma Khalid, Alastair G. Stewart, Stephen J. Matthews, Kay Grünewald, Carol V. Robinson