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.

Comprehensive Temporal Protein Dynamics during Postirradiation Recovery in Deinococcus radiodurans.

Deinococcus radiodurans (D. radiodurans) is an extremophile that can tolerate ionizing radiation, ultraviolet radiation, and oxidation. How D. radiodurans responds to and survives high levels of ionizing radiation is still not clear. In this study, we performed label-free proteomics to explore the proteome dynamics during postirradiation recovery (PIR). Surprisingly, proteins involved in translation were repressed during the initial hours of PIR. D. radiodurans also showed enhanced DNA repair and antioxidative response after 6 kGy of gamma irradiation. Moreover, proteins involved in sulfur metabolism and phenylalanine metabolism were enriched at 1 h and 12 h, respectively, indicating different energy and material needs during PIR. Furthermore, based on these findings, we proposed a novel model to elucidate the possible molecular mechanisms of robust radioresistance in D. radiodurans, which may serve as a reference for future radiation repair.

Confinement induces oxidative damage and synaptic dysfunction in mice.

A confined environment is an enclosed area where entry or exit is highly restricted, which is a risk factor for a work crew's mental health. Previous studies have shown that a crew is more susceptible to developing anxiety or depression in a confined environment. However, the underlying mechanism by which negative emotion is induced by confinement is not fully understood. Hence, in this study, mice were retained in a tube to simulate short-term confinement. The mice exhibited depressive-like behavior. Additionally, the levels of H2O2 and malondialdehyde in the prefrontal cortex were significantly increased in the confinement group. Furthermore, a label-free quantitative proteomic strategy was applied to analyze the abundance of proteins in the prefrontal cortex of mice. A total of 71 proteins were considered differentially abundant proteins among 3,023 identified proteins. Two differentially abundant proteins, superoxide dismutase [Mn] and syntaxin-1A, were also validated by a parallel reaction monitoring assay. Strikingly, the differentially abundant proteins were highly enriched in the respiratory chain, oxidative phosphorylation, and the synaptic vesicle cycle, which might lead to oxidative damage and synaptic dysfunction. The results of this study provide valuable information to better understand the mechanisms of depressive-like behavior induced by confined environments.

The tryptic peptides of hemoglobin for diagnosis of type 2 diabetes mellitus using label-free and standard-free LC-ESI-DMRM.

N-1-(deoxyfructosyl) valine of ß-hemoglobin, commonly referred to as HbA1c, is the "gold standard" for clinical detection of diabetes. Instead of quantifying the full-length HbA1c glycated protein, in the present study, we proposed the peptide-based strategy to quantify the depletion of the tryptic peptides of hemoglobin for the diagnosis of type 2 diabetes mellitus (T2DM). The peptides were discovered and validated as T2DM biomarkers by label-free LC-ESI-DMRM method without reference material. The glucose could react with hemoglobin's free amino group of N-terminus and ϵ-amino group of lysine residues and leave the modification on the hemoglobin tryptic peptides. Thus, there are two types of peptides in the hemoglobin: sensitive peptides and insensitive peptides to glucose due to the differential sensitivity of lysine residues to glycation. To discover two types of peptides of hemoglobin, we first developed the assay of liquid chromatography-electrospray ionization mass spectrometry coupled with dynamic multiple reaction monitoring. The protein coverage reaches 94.2%. Moreover, the hemoglobin was incubated with the 500 mmol/L glucose for 20 days, 40 days and 60 days in vitro to screen the sensitive peptides and insensitive peptides to glucose. A total of 14 sensitive peptides and 4 insensitive peptides were discovered. Furthermore, the LC-ESI-DMRM method was also utilized to validated the glucose-sensitive peptides by 40 clinical samples with healthy control individuals (n = 20) and type 2 diabetes mellitus patients (n = 20). Three putative sensitive peptides (LLGNVLVCVLAHHFGK, VVAGVANALAHKYH, LRVDPVNFK) from the hemoglobin showed excellent sensitivity and specificity based on receiver operating characteristic analysis and were verified as the promising biomarkers for the diagnosis of diabetes mellitus. And one peptide (LLVVYPWTQR) was found as glucose-insensitive peptide. Taken together, the findings of this study suggest that quantification of hemoglobin tryptic peptides using label-free and standard-free LC-ESI-DMRM is an alternative method for the diagnosis of T2DM, which could be combined with other MS-based blood biomarkers for diagnosis of multiple diseases in MS single shot.