• Peptide location fingerprinting reveals modification‐associated biomarker candidates of ageing in human tissue proteomes

      Ozols, Matiss; Eckersley, Alexander; orcid: 0000-0003-1602-4851; email: alexander.eckersley@manchester.ac.uk; Mellody, Kieran T.; Mallikarjun, Venkatesh; orcid: 0000-0002-0284-5205; Warwood, Stacey; O'Cualain, Ronan; Knight, David; Watson, Rachel E. B.; Griffiths, Christopher E. M.; Swift, Joe; et al. (2021-04-08)
      Abstract: Although dysfunctional protein homeostasis (proteostasis) is a key factor in many age‐related diseases, the untargeted identification of structurally modified proteins remains challenging. Peptide location fingerprinting is a proteomic analysis technique capable of identifying structural modification‐associated differences in mass spectrometry (MS) data sets of complex biological samples. A new webtool (Manchester Peptide Location Fingerprinter), applied to photoaged and intrinsically aged skin proteomes, can relatively quantify peptides and map statistically significant differences to regions within protein structures. New photoageing biomarker candidates were identified in multiple pathways including extracellular matrix organisation (collagens and proteoglycans), protein synthesis and folding (ribosomal proteins and TRiC complex subunits), cornification (keratins) and hemidesmosome assembly (plectin and integrin α6β4). Crucially, peptide location fingerprinting uniquely identified 120 protein biomarker candidates in the dermis and 71 in the epidermis which were modified as a consequence of photoageing but did not differ significantly in relative abundance (measured by MS1 ion intensity). By applying peptide location fingerprinting to published MS data sets, (identifying biomarker candidates including collagen V and versican in ageing tendon) we demonstrate the potential of the MPLF webtool for biomarker discovery.
    • Peptide Location Fingerprinting Reveals Tissue Region-Specific Differences in Protein Structures in an Ageing Human Organ

      Eckersley, Alexander; orcid: 0000-0003-1602-4851; email: alexander.eckersley@manchester.ac.uk; Ozols, Matiss; orcid: 0000-0001-5663-1053; email: matiss.ozols@manchester.ac.uk; Chen, Peikai; orcid: 0000-0003-1880-0893; email: pkchen@hku-szh.org; Tam, Vivian; email: vivtam@hku.hk; Hoyland, Judith A.; orcid: 0000-0003-4876-5208; email: judith.a.hoyland@manchester.ac.uk; Trafford, Andrew; orcid: 0000-0002-2770-445X; email: Andrew.W.Trafford@manchester.ac.uk; Chan, Danny; orcid: 0000-0003-3824-5778; email: chand@hku.hk; Sherratt, Michael J.; orcid: 0000-0003-4759-6617; email: michael.j.sherratt@manchester.ac.uk (MDPI, 2021-09-27)
      In ageing tissues, long-lived extracellular matrix (ECM) proteins are susceptible to the accumulation of structural damage due to diverse mechanisms including glycation, oxidation and protease cleavage. Peptide location fingerprinting (PLF) is a new mass spectrometry (MS) analysis technique capable of identifying proteins exhibiting structural differences in complex proteomes. PLF applied to published young and aged intervertebral disc (IVD) MS datasets (posterior, lateral and anterior regions of the annulus fibrosus) identified 268 proteins with age-associated structural differences. For several ECM assemblies (collagens I, II and V and aggrecan), these differences were markedly conserved between degeneration-prone (posterior and lateral) and -resistant (anterior) regions. Significant differences in peptide yields, observed within collagen I α2, collagen II α1 and collagen V α1, were located within their triple-helical regions and/or cleaved C-terminal propeptides, indicating potential accumulation of damage and impaired maintenance. Several proteins (collagen V α1, collagen II α1 and aggrecan) also exhibited tissue region (lateral)-specific differences in structure between aged and young samples, suggesting that some ageing mechanisms may act locally within tissues. This study not only reveals possible age-associated differences in ECM protein structures which are tissue-region specific, but also highlights the ability of PLF as a proteomic tool to aid in biomarker discovery.