How cells move gas

In the cover story of the September 10 issue of Science, scientists report the first high-resolution structural details of a protein that shuttles gas across cell membranes.The research team says the mechanism of ammonia permeation they find in bacteria is likely similar in eukaryotic cells. "The findings uncover a whole new biology of how this and orthologous proteins work, including Rh factors in humans," researcher Robert Stroud of the University of California, San Francisco, told The Scientist, because the shuttle proteins are genetically related to the structural components of the Rh blood group antigens of mammalian blood cells, which facilitate the transport of ammonia and carbon dioxide across eukaryotic cell membranes.Nobel laureate Peter Agre at Johns Hopkins University School of Medicine, Baltimore, who was not involved in this study, called the findings "a quantum leap forward" in the scientific understanding of gas transport. "It takes membrane gas permeation and goes from a low-level phenomenology to an atomic level of understanding in just one manuscript," Agre told The Scientist.While simple membrane bilayers are inherently moderately permeable to ammonia, the transport protein AmtB, present in the bacterial inner membrane between the cytoplasmic and periplasmic spaces, may serve to accelerate uptake at sites where ammonia diffusion is too slow for physiological needs. The mechanisms behind how gas transport proteins remained enigmatic until now. "It's very hard to uncover what the proteins do if they don't conduct ions, as the typical assays measure movement of electrical charge," Stroud said.To image AmtB, the researchers expressed paralogous genes for the protein from multiple bacterial species and prepared three-dimensional crystals of the proteins they encoded. The group then selected the crystal producing the highest resolution X-ray diffraction pattern for analysis. "The structure was solved remarkably quickly," Stroud said. "From having the perfect crystal to having the structure took under half an hour."They resolved AmtB's structure, with its 11 membrane-spanning helices, to 1.35 angstroms.AmtB "is a sticky, messy protein, and I would have predicted nobody would have achieved structure," said Agre, who coauthored an accompanying "Perspective" in Science with Mark Knepper of the National Heart, Lung, and Blood Institute in Bethesda, Maryland. "The resolution they've achieved is unprecedented" for a membrane protein, Agre said. "With the bacterial rhodopsins 1.5 angstroms was achieved. This work is as if the four-minute mile was suddenly run in three minutes." "The global importance of this study is that it gives hope the structures of many, many other integral membrane proteins can be solved. The success of Stroud and his group with such exquisite resolution bodes well for the approach they use. It looks to be powerful method and might find use for channels for sodium or urea or other solutes," Knepper, who was not involved in the study, told The Scientist.AmtB has the same trimeric structure when crystallized in both the absence and presence of ammonia, leading Stroud and colleagues to conclude it is a channel rather than a transporter that should have flexible elements involved in translocation of the substrate. At either ends of the pore, broader vestibules contain ammonia in equilibrium with ammonium. In AmtB's center midway through the membrane is a hydrophobic pore element roughly 20 angstroms long that allow ammonia to pass but not ammonium or other ions.The structure of Rh antigens to which AmtB is genetically related has long been pondered, and Agre and Knepper write in their "Perspective" that the trimeric structure of AmtB Stroud and colleagues uncovered now suggests how the three Rh polypeptides of red blood cells form the Rh antigen complex in the erythrocyte plasma membrane. They add that these findings could help predict how Rh-related proteins in the mammalian kidney collecting duct and liver mediate ammonia transport.

How cells move gas

Study reports high-resolution structure of protein that shuttles ammonia in bacteria

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