It is a rare event when a long-studied protein yields its structure to new experimental methods, and rarer still when that structure surprises the experts and intrigues even the casual observer. Such is the case for the recent model of a human voltage-dependent anion channel (VDAC-1), solved by nuclear magnetic resonance in the lab of Gerhard Wagner and reported in the Aug. 29 Science.
VDAC-1 allows passage of ions and small molecules including ATP across the mitochondrial outer membrane. It is also involved in controlling release of mitochondrial proteins that lead to programmed cell death. Wagner, the Elkan Blout professor of biological chemistry and molecular pharmacology at HMS, and his laboratory began working on the channel because, he said, “I realized that the most crucial events in apoptosis happen at the mitochondrial membrane.” Dysregulation of apoptosis can cause cancer and other disorders.
Human VDAC-1 joins 32 prokaryotic beta barrel membrane proteins whose structures have been solved. The beta strands are antiparallel, and all known beta barrels have an even number of strands. According to lead author Sebastian Hiller, a research fellow in Wagner’s lab, it was widely believed that all beta barrels—including eukaryotic ones—would display an even number of strands, since an odd number would make the terminal strands parallel, a less stable orientation.
The team studied recombinant VDAC-1 refolded in detergent balls called micelles, a system established by former graduate student Thomas Malia. Determining the protein’s fold involved assigning more than 600 nuclear Overhauser effects (NOEs) between atomic nuclei to define their spatial proximity and therefore the overall topology of the protein. Initially, Hiller and Wagner thought VDAC-1 fit the mold with an even number of strands, but suddenly, each independently realized that the data were telling them there were 19 strands. “Gerhard said, ‘It must have 19,’” Hiller recalled, “and I had just found two new NOEs,” which convinced them.
One striking feature of the structure is its ability to account for VDAC-1’s ion preferences: one negative and two positive patches are seen, and VDAC-1 displays a 2:1 preference for anions. An experiment with detergent carrying a paramagnetic spin label identified a belt of amino acid residues contacting detergent. Interaction sites with binding partners such as the anti-apoptotic protein Bcl-x(L) were also observed.
Hiller and Wagner expect that the topology in the micelles is the same in lipid bilayers. “We are not aware of any structure where a membrane protein adopts different structures in different environments,” Wagner said. Nonetheless, they are keen to take on the challenge of studying the protein in a bilayer.
Despite VDAC-1’s odd number of beta strands, both termini still lie on the same side of the channel, thanks to a long N-terminal tail that reaches through it. “We think this part is crucial for gating,” Wagner said. The group has begun a collaboration to study by computational modeling how gating is accomplished.
In fact, Hiller said, “we have a list of 30 or 40 things that I would like to do.”
Conflict Disclosure: The authors declare no conflicts of interest.
Funding Sources: The National Institutes of Health; the Swiss National Science Foundation; the Wenner-Gren Foundation, Stockholm; and the Ludwig Foundation for Cancer Research
