Large‐scale protein identification of human urine proteome by multi‐dimensional LC and MS/MS

Abstract

Urine is a human specimen that is easily obtained non-invasively for clinical diagnosis. We attempted to enhance the resolution of current human urine proteomes and construct a comprehensive reference database for advanced studies, such as the discovery of biomarkers for renal diseases. Multi-dimensional LC-MS/MS was coupled with de novo sequencing and database matching. The proposed approach improved the identification of not only the proteins, but also the post-translational sites of urinary proteins. We identified 165, 200 and 259 unique gene products in the urine proteomes from males, females and pregnant women, respectively. When all of the results were combined and the redundancies removed, a total of 1095 distinct peptides were identified. Of these, 1016 peptides were associated with 334 unique gene products. In this study, over 100 gene products, including some disease-related proteins, were detected in urine for the first time by proteomic approaches. Various proteins with novel post-translational hydroxylation were identified using the MASCOT program and de novo sequencing. All proteins with peptide information were summarized into a comprehensive urine protein database. We believe that this comprehensive urine proteome database will assist in the identification of urinary proteins/polypeptides whose spectra are difficult to interpret in the discovery of urinary biomarkers.

Contextualised urinary biomarker analysis facilitates diagnosis of paediatric obstructive sleep apnoea

“…Indeed, gene ontology analysis of the detected urinary proteome identified significant enrichments in functional annotations (Fig. 1e) consistent with previous proteomics analyses of urine (33, 34). …”

Discovery of novel candidate urinary protein biomarkers for prostate cancer in a multiethnic cohort of South African patients via label‐free mass spectrometry

“…Studies from America, Europe, and parts of Asia have previously described and characterized a catalog of normal and disease‐related human urinary proteomes , identifying from 124 to 1823 urinary proteins. Two landmark papers have recently described a complete draft of the human proteome, using several adult and fetal human tissues, body fluids, and cell lines to identify 17 294–18 097 of the ca.…”

Multiplexed quantification of 63 proteins in human urine by multiple reaction monitoring-based mass spectrometry for discovery of potential bladder cancer biomarkers

“…The profiles of the human urinary proteome, particularly those proteins that are associated with kidney function and the urinary system, vary with disease progression or drug treatment. Thus, interest in exploring the urinary proteome has broadened the search for new biomarkers, as well as disease etiology studies [7,13,[41][42][43][44]. Protein biomarker discovery in urine has gained prominence in recent years, and levels of specific proteins in urine have been found to increase or decrease during disease in numerous studies, including kidney disease, bladder disease, and cancers [7,[45][46][47].…”

“…With the incorporation of multidimensional fractionation, immuno-enrichment or depletion, the cataloging of proteins has been improved [12][13][14][15][16]. However, because of the limited sampling rate of mass spectrometers using untargeted analyses, and because of technical variations in multidimensional proteomics studies, conventional bottom-up proteomics has only limited reproducibility for protein quantitation purposes.…”