Revealing sex-specific changes across protein structure in the aging bone extracellular matrix

The process of aging is an integral but complex component of life that has been intensely studied for decades, from the molecular level to whole organisms. At the tissue level, bone is one of the most difficult to study due to its composite nature of inorganic and organic phases, but advancements in proteomics are enhancing our understanding of the latter, improving our understanding of skeletal aging through identifying temporal changes across the bone proteome. The relative longevity of extracellular matrix (ECM) proteins can make them more susceptible to accumulating damage modifications over time. In addition, their informational density, including their 2D and 3D structure, protein folding, post translational modifications and proteomic composition, as well as their functional importance, has made them a target of interest in the study of aging and medical conditions such as osteoporosis and arthritis. ECM proteins are also increasingly utilised in forensic science for determining biological sex/age because of their longevity. Following recent developments in peptide location fingerprinting methods that improve capabilities of identifying regional changes in protein structure, this study aimed to identify age-associated regional changes along protein structures in <i>Rattus norvegicus</i> from whole limb LC-MC/MS data. Regional changes in protein structure were identified in a variety of collagenous and non-collagenous ECM proteins, providing evidence for increased remodeling in juvenile rats and a reduced ability in adult rats, alongside damage accumulation in the adult ECM. This research highlights the importance of fibrillar collagen remodeling but is also indicative of potential new roles for osteopontin, thrombin, apolipoproteins and wider ECM regulators such as cartilage oligomeric matrix protein. This demonstrates, in this case, the utility of peptide location fingerprinting as a screening tool to identify biomarker candidates of bone aging between juvenile and adult rats.