Discovery and validation of glucose-sensitive peptide biomarkers from human serum albumin to diagnose type 2 diabetes mellitus

Glycated albumin (GA), which represents the global glycation level of albumin, has emerged as a biomarker for diagnosing prediabetes and diabetes. In our previous study, we developed a peptide-based strategy and found three putative peptide biomarkers from the tryptic peptides of GA to diagnose type 2 diabetes mellitus (T2DM). However, the trypsin cleavage sites at the carboxyl side of lysine (K) and arginine (R) are consistent with the nonenzymatic glycation modification site residues, which considerably increases the number of missed cleavage sites and half-cleaved peptides. To solve this problem, the endoproteinase Glu-C was used to digest GA from human serum to screen putative peptides to diagnose T2DM. In the discovery phase, we found eighteen and fifteen glucose-sensitive peptides from purified albumin and human serum incubated with 13C glucose in vitro, respectively. In the validation phase, eight glucose-sensitive peptides were screened and validated in 72 clinical samples (28 healthy controls and 44 patients with diabetes) using label-free LC-ESI-MRM. Three putative sensitive peptides (VAHRFKDLGEE, FKPLVEEPQNLIKQNCE and NQDSISSKLKE) from albumin exhibited good specificity and sensitivity based on receiver operating characteristic analysis. In summary, three peptides were found as promising biomarkers for the diagnosis and assessment of T2DM based on mass spectrometry. [Display omitted] •We quantify the depletion of Glu-C digested peptides of albumin to screen the putative peptide biomarkers of T2DM.•Glucose-sensitive peptides are discovered by albumin glycation and serum glycation models and validated by clinical samples.•Three glucose-sensitive peptides are found as promising biomarkers for diagnosis of T2DM.•High-abundant peptides are easier to detect and more suitable for clinical diagnosis of T2DM by mass spectrometry.