Abstract

Phosphorylation plays a critical role in regulating protein function and thus influences a vast spectrum of cellular processes. With the advent of modern bioanalytical technologies, examination of protein phosphorylation on a global scale has become one of the major research areas. Phosphoproteins are found in biological fluids and interrogation of the phosphoproteome in biological fluids presents an exciting opportunity for discoveries that hold great potential for novel mechanistic insights into protein function in health and disease, and for translation to improved diagnostic and therapeutic approaches for the clinical setting. This review focuses on phosphoproteome discovery in selected human biological fluids: serum/plasma, urine, cerebrospinal fluid, saliva, and bronchoalveolar lavage fluid. Bioanalytical workflows pertinent to phosphoproteomics of biological fluids are discussed with emphasis on mass spectrometry-based approaches, and summaries of studies on phosphoproteome discovery in major fluids are presented.

Highlights

  • Phosphorylation is a common post-translational modification of proteins that involves the reversible attachment of phosphate groups to the side chains of specific amino acids.O-phosphorylation occurs most commonly on serine (Ser) and threonine (Thr) residues; a small fraction of phosphorylation is present on tyrosines (Tyr) [1]

  • Phosphoproteomics interrogation of major biological fluids provides an attractive opportunity to gain expanded, unique scientific knowledge complementary to global-scale profiling of protein expression. This unique knowledge on protein phosphorylation holds the promise of new mechanistic insights into protein function in health and disease, and the potential to be translated into clinical applications for improved patient care outcomes

  • There are a handful of groups who have published in this arena, and the pace of moving beyond initial phosphoproteome discovery has generally been slow

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Summary

Introduction

Phosphorylation is a common post-translational modification of proteins that involves the reversible attachment of phosphate groups to the side chains of specific amino acids. Serum/plasma reflects the status of distant tissues, which collectively contribute to the overall (phospho)proteome profile Proximal fluids such as cerebrospinal fluid (CSF) or bronchoalveolar lavage fluid (BAL) reflect the health/disease processes of the particular organ(s); these fluids are more likely to contain higher concentrations of organ-specific marker proteins and provide a more direct molecular readout of the local milieu from which they originate. As a common approach adopted by the research groups engaged in biological fluid phosphoproteomics, the first stage of the research centers on qualitative phosphoproteome discovery Such qualitative phosphoproteome survey aims to provide description of the catalog of phosphoproteins and the exact assignment of the phosphorylation sites. In reviewing the bioanalytical strategies, we focus mainly on features pertinent to published biological fluid phosphoproteomics studies, with some discussion of new developments for potential inclusion in future workflows. In the second part of the review, we present synopses of phosphoproteome studies in individual fluids that have been published to date, and give our opinions on future directions in this field

General Workflow Characteristics
Protein Processing
Protein Digestion
Separation of Peptide Mixtures
Phosphopeptide Enrichment
Bioinformatics
Applications to Biofluid Phosphoproteome Characterization
Cerebrospinal Fluid
Saliva
Bronchoalveolar Lavage Fluid
Concluding Remarks
Findings
Methods
Full Text
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