Chromatography combined with database search strategy identifies hard-to-find heme proteins
“Results: Heme c is an important iron-containing post-translational modification found in many proteins. It plays an important role in respiration, metal reduction, and nitrogen fixation, especially anaerobic respiration of environmental microbes. Such bacteria and their c-type cytochromes are studied extensively because of their potential use in bioremediation, microbial fuel cells, and electrosynthesis of valuable biomaterials.
Until recently, these modifications were hard to find using traditional proteomic methods. Scientists at Pacific Northwest National Laboratory combined a heme c tag protein affinity purification strategy called histidine affinity chromatography (HAC) with enhanced database searching. This combination confidently identified heme c peptides in liquid chromatography-tandem mass spectrometry (LC-MS/MS) experiments-by as much as 100-fold in some cases.”
Why It Matters: Iron is a critical part of many biological processes; however, it is often not biologically available or it can be toxic in high quantities. So, biological systems have developed intricate methods to use and store iron. Many environmentally important microbes and microbial communities are rich in c-type cytochromes. Combining HAC and data analysis tailored to the unique properties of heme c peptides should enable more detailed study of the role of c-type cytochromes in these microbes and microbial communities.
‘Several proteomics studies have analyzed the expression of c-type cytochromes under various conditions,’ said PNNL postdoctoral researcher Dr. Eric Merkley, and lead author of a paper that appeared in the Journal of Proteome Research. ‘A shared feature of these studies is that the cytochrome-rich fractions, the cell envelope or extracellular polymeric substance, were purified and explicitly analyzed to efficiently detect cytochromes. Analyses of large-scale proteomics datasets have typically suggested that c-type cytochromes, particularly the heme c peptides, are under-represented.'”
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Pacific Northwest National Laboratory (PNNL) is one of the United States Department of Energy National Laboratories, managed by the Department of Energy’s Office of Science. The main campus of the laboratory is in Richland, Washington.
PNNL scientists conduct basic and applied research and development to strengthen U.S. scientific foundations for fundamental research and innovation; prevent and counter acts of terrorism through applied research in information analysis, cyber security, and the nonproliferation of weapons of mass destruction; increase the U.S. energy capacity and reduce dependence on imported oil; and reduce the effects of human activity on the environment. PNNL has been operated by Battelle Memorial Institute since 1965.
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