For fifty years, biochemists suspected that proteins shed their protective water shells when their environment turned acidic—but nobody had ever actually seen it happen. Now researchers at Martin Luther University Halle-Wittenberg have done something remarkable: using cryo-EM technology, they've watched thousands of individual water molecules abandon a protein as acidity increases, finally confirming a theory that has lingered unanswered since 1974.
The discovery matters because proteins are cellular machines that drive everything from growth to metabolism in living organisms. Their job depends entirely on maintaining the right shape, and that shape is stabilized by a thin skin of water molecules that clings to their surface. Until now, the relationship between a protein's water shell, acidity levels, and overall structure remained largely mysterious—a gap in fundamental biochemistry that researchers had puzzled over for decades.
Professor Panagiotis Kastritis and his team at MLU tackled this by studying apoferritin, a protein that stores iron in cells. They used cryogenic electron microscopy, a technique that flash-freezes proteins and bombards them with an electron beam to generate impossibly detailed images. They observed apoferritin at seven different pH levels, ranging from slightly alkaline at 9.0 to quite acidic at 3.5, meticulously mapping thousands of water molecules and tracking how each one behaved as the environment changed.
What emerged was startling precision. For every single unit by which pH decreases—meaning the environment becomes more acidic—the protein sheds approximately 100 water molecules. But the process isn't random. Dr. Ioannis Skalidis, who completed his Ph.D. at MLU and now works at Utrecht University, was surprised to discover clear rules governing which water molecules stick around and which depart. "Certain amino acids bind water, while others release it," he explains. "We hadn't expected to see this."
Two specific amino acids, glutamate and aspartate, are the first to abandon their water molecules when acidity rises. Other amino acids stubbornly hold onto their water regardless of pH shifts, maintaining a stable interior core made up of roughly 40 percent of the total water molecules. This inner sanctuary appears to remain unchanged no matter what the external conditions are.
The team uncovered an additional surprise: as pH drops and the water shell shifts, the iron ions bound within the protein also move, gradually sliding away from their binding sites. This reveals a structural mechanism by which increasing acidity can trigger the release of metal ions from proteins—a process fundamental to how cells regulate iron levels.
"We are the first group to succeed in providing this evidence at this level of mechanistic detail," says Dr. Farzad Hamdi from MLU. The findings, published in the Proceedings of the National Academy of Sciences in 2026, open a new door for protein engineering. If these rules apply broadly across different proteins—a question future studies will need to answer—researchers could design proteins that are more stable or better able to tolerate varying pH levels. Such advances could improve industrial enzymes and protein-based drug delivery systems, bringing decades-old theory into practical application.