Protein phosphorylation helps control most processes in cells. Phosphorylation most often occurs at serine, threonine, or tyrosine residues in proteins. Histidine residues also can be phosphorylated, but biologists have assumed this is rare. A new study suggests that histidine phosphorylation might be more common than previously thought.
Albert J. R. Heck, Simone Lemeer, and coworkers at the University of Utrecht have developed a new proteomics approach for analyzing histidine phosphorylation (Nat. Methods 2018, DOI: 10.1038/nmeth.4580). They use Fe3+-immobilized metal affinity chromatography to enrich peptides with phosphorylated histidines in samples and then analyze the peptides by liquid chromatography-tandem mass spectrometry. By applying milder conditions than are typically used in phosphopeptide enrichments, the team can prevent hydrolysis of the unstable histidine phosphorylation.
The researchers demonstrated the power of the method by studying protein histidine phosphorylation in Escherichia coli. They detected 2,129 phosphorylation sites, which is about 10 times as many as seen with established analytical methods. Of those sites, 246 were phosphohistidine sites, occurring on 173 bacterial proteins. The researchers were surprised to find that histidine phosphorylation represented about 10% of the overall E. coli phosphoproteome, making it about as abundant and frequent as phosphorylation on threonine or tyrosine residues.
The researchers think that the new method “could help answer long-standing questions about whether histidine phosphorylation also plays a major biological role in higher organisms,” including humans.