Peroxide-Induced Damage to Plasminogen Molecules

Doklady Biochemistry and Biophysics - Plasminogen is a zymogenic form of plasmin, an enzyme that plays a fundamental role in the dissolution of fibrin clots as well as in many other physiological...     

Determination of proteomic and metabolic composition of exhaled breath condensate of newborns

Here, the possibility of proteomic and metabolomic analysis of the composition of exhaled breath condensate of neonates with respiratory support. The developed method allows non-invasive collecting sufficient amount of the material for identification of disease-specific biomarkers. Samples were collected by using a condensing device that was incorporated into the ventilation system. The collected condensate was analyzed by liquid chromatography coupled with high resolution mass spectrometry and tandem mass spectrometry. The isolated substances were identified with a use of databases for proteins and metabolites. As a result, a number of compounds that compose the exhaled breath condensate was determined and can be considered as possible biomarkers of newborn diseases or stage of development.     

Preconcentration of organic solutes in urine by bubble bursting

Introduction

The chemical sensitivity of urine metabolomics analysis is greatly compromised due to the large amounts of inorganic salts in urine (NaCl, KCl), which are detrimental to analytical instrumentation, e.g. chromatographic columns or mass spectrometers. Traditional desalting approaches applied to urine pretreatment suffer from the chemical losses, which reduce the information depth of analysis.

Objectives

We aimed to test a simple approach for the simultaneous preconcentration and desalting of organic solutes in urine based on the collection of induced bursting bubble aerosols above the surface of urine samples.

Method

Bursting bubbles were generated at ambient conditions by feeding gas through an air diffuser at the bottom of diluted (200 times in ultrapure water) urine solution (50–500 mL). Collected aerosols were analyzed by the direct-infusion electrospray ionization mass spectrometry (ESI–MS).

Results

The simultaneous preconcentration (ca. 6–12 fold) and desalting (ca. six–tenfold) of organic solutes in urine was achieved by the bursting bubble sample pretreatment, which allowed ca. three-times higher number of identified urine metabolites by high-resolution MS analysis. No chemical losses due to bubbling were observed. The increased degree of MS data clustering was demonstrated on the principal component analysis of data sets from the urine of healthy people and from the urine people with renal insufficiency. At least ten times higher sensitivity of trace drug detection in urine was demonstrated for clenbuterol and salbutamol.

Conclusion

Our results indicate the high versatility of bubble bursting as a simple pretreatment approach to enhance the chemical depth and sensitivity of urine analysis. The approach could be attractive for personalized medicine as well as for the diagnostics of renal disorders of different etiology (diabetic nephropathy, chronic renal failure, transplant-associated complications, oncological disorders).

Graphical Abstract

Urine desalting and preconcentration in bursting bubbles.

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Accurate mass tag retention time database for urine proteome analysis by chromatography-mass spectrometry

Information about peptides and proteins in urine can be used to search for biomarkers of early stages of various diseases. The main technology currently used for identification of peptides and proteins is tandem mass spectrometry, in which peptides are identified by mass spectra of their fragmentation products. However, the presence of the fragmentation stage decreases sensitivity of analysis and increases its duration. We have developed a method for identification of human urinary proteins and peptides. This method based on the accurate mass and time tag (AMT) method does not use tandem mass spectrometry. The database of AMT tags containing more than 1381 AMT tags of peptides has been constructed. The software for database filling with AMT tags, normalizing the chromatograms, database application for identification of proteins and peptides, and their quantitative estimation has been developed. The new procedures for peptide identification by tandem mass spectra and the AMT tag database are proposed. The paper also lists novel proteins that have been identified in human urine for the first time.     

Proteomic Markers and Early Prediction of Alzheimer’s Disease

Biochemistry (Moscow) - Alzheimer’s disease (AD) is the most common socially significant neurodegenerative pathology, which currently affects more than 30 million elderly people worldwide....     

Exogenous proteins in exhaled human breath condensate

In the course of analysis of protein composition of exhaled breath to diagnose diseases of the respiratory system the problem is raised to distinguish between proteins, expressed in lung tissues and in respiratory tract (endogenous) and those that got into the respiratory system from the ambient air in the process of respiration (exogenous). In this work, an attempt is made to estimate the constitution of exogenous proteins in exhaled air with mass spectrometry and nanoflow high performance liquid chromatography (nano-HPLC). Six months’ indoors isolation of healthy donors with air being cleaned of dust leads to the removal from the spectrum of exhaled proteins of some keratins that are therefore considered to be exogenous. Nonkeratin proteins may also circulate between ambient air and human airways, but their concentration appears to be significantly lower than keratin concentrations (especially than the epidermis keratin). Among nonkeratins, dermicidin seems to be the most significant exogenous protein of the exhaled air. Conclusions concerning the diagnostic value of exhaled proteins can be made only after careful comparison of results of quantitative and qualitative analyses of their normal and pathological composition for a statistically significant sample of donors.     

Potassium Ions are More Effective than Sodium Ions in Salt Induced Peptide Formation

Prebiotic peptide formation under aqueous conditions in the presence of metal ions is one of the plausible triggers of the emergence of life. The salt-induced peptide formation reaction has been suggested as being prebiotically relevant and was examined for the formation of peptides in NaCl solutions. In previous work we have argued that the first protocell could have emerged in KCl solution. Using HPLC-MS/MS analysis, we found that K⁺ is more than an order of magnitude more effective in the L-glutamic acid oligomerization with 1,1'-carbonyldiimidazole in aqueous solutions than the same concentration of Na⁺, which is consistent with the diffusion theory calculations. We anticipate that prebiotic peptides could have formed with K⁺ as the driving force, not Na⁺, as commonly believed.     

Modification of the Urine Proteome in Healthy Human during 21-day Bed Rest

Mass spectrometry–based proteomics was employed to analyze urine from eight healthy volunteers during a 21-day bed rest (BR) study. The analysis included trypsinolysis in solution prior to liquid chromatography and tandem mass spectrometry (LC-MS/MS), and spectrum processing using the bioinformatic tools. Relying on 221 IPI indices with scores from 24 to 1700, 169 different proteins were identified. Molecular functions, biological processes, and cell components as the loci of certain protein functioning were determined with the help of UniProt-GOA. Associative interactions networks were constructed using BiNGO. There were 14 proteins identified that were functional in the cardiovascular system mostly. They were annotated, and the dynamics of their occurrence throughout the experiment was considered. Grounding on the biological functions of these proteins and an assumption of eligible activation of different biological processes during BR was made.     

Oxidation-induced modification of the fibrinogen polypeptide chains

By using the mass-spectrometry method, the oxidative modifications of the fibrinogen Aα, Bβ, and γ polypeptide chains induced by its oxidation have been studied. The αC-region has been proven to be the most vulnerable target for the oxidizer (ozone) as compared with the other structural elements of the Aα chain. The Bβ chain mapping shows that the oxidative sites are localized within all the structural elements of the chain in which the β-nodule exhibits high susceptibility to oxidation. The γ chains are the least vulnerable to the oxidizer action. The results obtained demonstrate convincingly that the self-assembly centers dealing with interactions of knob “A”: hole “a” are not involved in oxidative modification. It is concluded that the numerous oxidative sites revealed are mainly responsible for inhibiting lateral aggregation of protofibrils. The part of amino acid residues subjected to oxidation is supposed to carry out the antioxidant function.     

Estimation of phosphorylation level of amyloid-beta isolated from human blood plasma: Ultrahigh-resolution mass spectrometry

Recently, amyloid-beta (Aβ) phosphorylation at position 8 has been shown to be associated with pathogenesis of Alzheimer’s disease. Since the modification occurs in the key fragment of the metal-binding domain of Aβ and should seriously affect the interaction of pS8-Aβ with zinc ions, this isoform may be a potential precursor of pathogenic oligomer forms of Aβ. Hence the level of pS8-Aβ in human biological fluids (blood, urine, cerebrospinal fluid) may reflect various stages of pathogenesis of the Alzheimer’s disease. The aim of the work was to develop a prototype of an analytical method for quantitative determination of the level of pS8-Aβ isoform in binary mixtures with native Aβ in order to further use it to estimate the levels of phosphorylated amyloid-beta in blood plasma samples of patients diagnosed with Alzheimer’s disease.