title: Investigations into the Use of a Protein Sensor Assay for Metabolite Analysis.
abstract: Rapid and definitive classification of biological samples has application in industrial, agricultural, and clinical settings. Considerable effort has been given to analytical methods to address such applications over the past 50 years, with the majority of successful solutions focusing on a single molecular target. However, in many cases, a single or even a few features are insufficient for accurate characterization or classification. Serum albumin (SA) proteins are a class of cargo-carrying proteins in blood that have evolved to transport a wide variety of metabolites and peptides in mammals. These proteins have up to seven binding sites which communicate allosterically to orchestrate a complex pick-up and delivery system involving a large number of different molecules at any time. The ability of SA proteins to bind multiple molecular species in a sophisticated manner inspired the development of assays to differentiate complex biological solutions. The combination of SA and high-resolution liquid chromatography mass spectrometry (LC-MS) is showing exciting promise as a protein sensor assay (PSA) for classification of complex biological samples. In this study, the PSA has been applied to cells undergoing and recovering from mild oxidative stress. Analysis using traditional LC-MS-based metabolomics failed to differentiate samples into treatment or temporal groups, whereas samples first treated with the PSA were cleanly classified into both correct treatment and temporal groups. The success of the PSA could be attributed to selective binding of metabolites, leading to a reduction in sample complexity and a general reduction in chemical noise. Metabolites important to successful sample classification were often enriched by 100-fold or more yet displayed a wide range of affinities for SA. The end result of PSA treatment is better classification of samples with a reduction in the number of features seen overall. Together, these results demonstrate how the use of a protein-based assay before LC-MS analysis can greatly improve separation and lead to more accurate and successful tracking of the metabolic state in an organism, suggesting potential application in a wide range of fields.
authors.affiliation: Department of Chemistry and Biochemistry, Montana State University, Bozeman, MT, 59717, USA.
authors.forename: Timothy
authors.initials: T
authors.lastname: Hamerly
authors.affiliation: Department of Chemistry and Biochemistry, Montana State University, Bozeman, MT, 59717, USA. bbothner@chemistry.montana.edu.
authors.forename: Brian
authors.initials: B
authors.lastname: Bothner
mesh_terms: D002853:Chromatography, Liquid; D013058:Mass Spectrometry; D055432:Metabolomics / Q000379:methods*; D011506:Proteins / Q000378:metabolism*
chemical_list: D011506:Proteins
keywords: Biomarker; Bovine serum albumin; LC-MS; Metabolomics; Oxidative stress; Protein sensor