An investigation into the human serum “interactome”

TLDR

Human serum contains proteins from nearly all body tissues, and its proteome reflects disease states, but the vast dynamic range and routine depletion of abundant proteins hinder complete characterization and may also remove associated low‑abundant proteins. In this study, immunoprecipitation was combined with μRPLC‑MS/MS to investigate low‑molecular‑weight proteins/peptides that associate with the most abundant serum proteins. Targeted isolation of abundant serum proteins by immunoprecipitation enriches associated proteins/peptides, which are then identified by μRPLC‑MS/MS. The approach identified 210 proteins, of which 73 % were novel to low‑molecular‑weight studies and 67 % were novel to whole‑serum proteome studies.

Abstract

Abstract The protein content of human serum is composed of a millieu of proteins from almost every type of cell and tissue within the body. The serum proteome has been shown to contain information that directly reflects pathophysiological states and represents an invaluable source of diagnostic information for a variety of different diseases. Unfortunately, the dynamic range of protein abundance, ranging from ≫ mg/mL level to ≪ pg/mL level, renders complete characterization of this proteome nearly impossible with current analytical methods. To study low‐abundance proteins, which have potential value for clinical diagnosis, the high‐abundant species, such as immunoglobulins and albumin, are generally eliminated as the first step in many analytical protocols. This step, however, is hypothesized to concomitantly remove proteins/peptides associated with the high‐abundant proteins targeted for depletion. In this study, immunoprecipitation was combined with microcapillary reversed‐phase liquid chromatography (μRPLC) coupled on‐line with tandem mass spectrometry (MS/MS) to investigate the low‐molecular‐weight proteins/peptides that associate with the most abundant species in serum. By this targeted isolation of select highly abundant serum proteins, the associated proteins/peptides can be enriched and effectively identified by μRPLC‐MS/MS. Among the 210 proteins identified, 73% and 67% were not found in previous studies of the low‐molecular‐weight or whole‐serum proteome, respectively.