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Standards for Quantitative Metalloproteomic Analysis Using Size Exclusion ICP-MS
Published on: April 13, 2016
Multiple forms of selenoprotein P in a candidate human plasma standard reference material
Guillaume Ballihaut1, Lisa E Kilpatrick, Eric L Kilpatrick
1National Institute of Standards and Technology, Analytical Chemistry Division, Hollings Marine Laboratory, 331 Fort Johnson Road, Charleston, SC 29412, USA. guillaume.ballihaut@noaa.gov
Metallomics : Integrated Biometal Science
|May 4, 2012
Summary
This study identifies three selenoprotein P (SePP) isoforms in human plasma using LC-MS/MS, with SePP49 and SePP57 containing selenium. SePP45 is a truncated form, and SePP may interact with GPx3 in vivo.
Area of Science:
- Biochemistry
- Proteomics
- Trace element metabolism
Background:
- Selenoprotein P (SePP) transports selenium (Se) and is a key nutritional marker.
- Previous Western blot studies identified SePP isoforms (51 kDa, 61 kDa), but their structure and Se content were unclear.
- The 51 kDa isoform was suspected to be a purification artifact.
Purpose of the Study:
- To characterize SePP isoforms in human plasma using advanced analytical techniques.
- To determine the selenium content and sequence integrity of identified SePP isoforms.
- To investigate potential interactions between SePP and other proteins in vivo.
Main Methods:
- Immunoprecipitation of SePP from human plasma Standard Reference Material.
- Liquid chromatography-tandem mass spectrometry (LC-MS/MS) for isoform identification and sequencing.
- Gel electrophoresis coupled with laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) for selenium detection.
Main Results:
- Three SePP isoforms were identified at 45 kDa, 49 kDa, and 57 kDa (SePP45, SePP49, SePP57).
- SePP49 and SePP57 contained selenium and yielded selenopeptides, confirming their integrity.
- SePP45 lacked selenium and relevant peptides, indicating it's a truncated form. A 23 kDa band contained selenium and glutathione peroxidase 3 (GPx3).
Conclusions:
- Human plasma contains three distinct SePP isoforms, SePP45, SePP49, and SePP57.
- SePP45 is a transcriptionally truncated isoform, while SePP49 and SePP57 are selenium-containing.
- Preliminary evidence suggests in vivo interaction between SePP and GPx3.
