Laccase-Catalyzed Heterocoupling of Dihydroquercetin and p-Aminobenzoic Acid: Effect of the Reaction Product on Cultured Cells.

Derivatization of the natural flavonoid dihydroquercetin with p-aminobenzoic acid was carried out in an ethyl acetate/citric buffer biphasic system using laccase from the fungus Trametes hirsuta. The main reaction product yield was ~68 mol %. The product was characterized by 1H NMR, 13C NMR, and liquid chromatography-mass spectroscopy, and its structure was elucidated. The reaction product affected viability of cultured human rhabdomyosarcoma cells (RD cell line) in a dose-dependent manner and, therefore, can be of interest to pharmaceutical industry.

[The application of proteomic technologies for the analysis of muscle proteins of farm animals used in the meat industry (review)].

The review briefly summarizes the data on the development of proteomic technologies that became actively used in studies of the muscular proteins of farm animals used in the meat industry in 2006-2013. It has been noted that the main research trends are connected with the detection of changes in muscle proteins during post-mortem autolysis and the search for species-specific and other protein biomarkers. Particular publications regarding the development of methods based on proteomic technologies for monitoring the state of muscle proteins are considered. According to the analyzed data, we can conclude that the field is promising for the solution of a number of pressing problems in.applied biochemistry.

AGR2, ERp57/GRP58, and some other human protein disulfide isomerases.

This review considers the major features of human proteins AGR2 and ERp57/GRP58 and of other members of the protein disulfide isomerase (PDI) family. The ability of both AGR2 and ERp57/GRP58 to catalyze the formation of disulfide bonds in proteins is the parameter most important for assigning them to a PDI family. Moreover, these proteins and also other members of the PDI family have specific structural features (thioredoxin-like domains, special C-terminal motifs characteristic for proteins localized in the endoplasmic reticulum, etc.) that are necessary for their assignment to a PDI family. Data demonstrating the role of these two proteins in carcinogenesis are analyzed. Special attention is given to data indicating the presence of biomarker features in AGR2 and ERp57/GRP58. It is now thought that there is sufficient reason for studies of AGR2 and ERp57/GRP58 for possible use of these proteins in diagnosis of tumors. There are also prospects for studies on AGR2 and ERp57/GRP58 leading to developments in chemotherapy. Thus, we suppose that further studies on different members of the PDI family using modern postgenomic technologies will broaden current concepts about functions of these proteins, and this will be helpful for solution of urgent biomedical problems.

[Study of Dj-1 protein in tissue specimens, cultured cells and serum of prostate cancer patients].

Two isoforms of Dj-1 protein were identified using a proteomic study in tissue specimens from two groups of patients with confirmed benign prostate hyperplasia (BPH) and prostate cancer (PCa). Dj-1 was also found in the cell lines PC-3, DU-145, LNCaP, BPH-1, and the lowest level of Dj-1 was found in BPH-1. Immunochemical study (ELISA) of serum levels of Dj-1, Bcl-2, IGF-1 and IGFBP-3 proteins revealed statistically significant distinctions between two groups of patients (p=0,004, Mann-Whitney test) only for Dj-1. Taken together, these data suggest that Dj-1 protein is a perspective biomarker candidate for PCa.

[The study of adaptation mechanisms of Yarrowia lipolytica yeast to alkaline conditions by means of proteomics].

Using proteomic technologies--two-dimensional electrophoresis in denaturing conditions in combination with mass spectroscopy of MALDI-TOF proteins--we demonstrated, for the first time, that the most noticeable alteration of protein composition of a Yarrowia lipolytica cell during adaptation to alkaline conditions was an increase of mitochondrial proteins relatively to proteins of cytoplasm.

[A study of protein profile changes in differentiating human myoblasts].

The changes in the protein profile in cultured human myoblasts after induction of differentiation was studied by proteomic techniques (a combination of O'Farrell two-dimensional electrophoresis and subsequent protein identification by MALDI-TOF MS and MS/MS analyses). Forty-one proteins have been identified, 25 of which were present in both proliferating and differentiating myoblasts, which allows them to be considered as myoblast housekeeping proteins. The changes in the distribution of some isoforms of tropomyosins, S100 proteins, cofilin, etc. have been revealed. The possible role of these changes in the cell protein profile in the realization of the program of skeletal muscle cell differentiation is discussed.

[Determination of "amino acids conflicts" and amino acids substitutions in primary structures of 41 human proteins by use the proteomic technologies].

Proteomic studies of some human tissues and organs (skeletal muscles, myometrium, motor zone of the brain, prostate), and also cultivated myoblasts revealed 41 of 300 identified proteins, in which the present of certain variants of amino acids ("conflicts") was recognized at several positions. Among the 93 registered amino acids "conflicts", seven cases represented the results of the protein polymorphisms caused by corresponding substitution of individual amino acid. Moreover, among prostate proteins the proteomic analysis revealed two isoforms of prostate-specific antigen, formed due to alternative splicing. Thus, our results have shown, that proteomic technologies allow to specify effectively the features of primary structures and to characterize various kinds of polymorphism in many human proteins.

[Age-related changes of albumin and actin of human myocardium].

In the human myocardium tissue, proteomic technologies revealed the products of 2 genes (alpha-actin and albumin), existing as fragments; their appearance and increased contents correlated with age. The age-related variants differ from the mature forms by the absence of N-terminal fragments of the amino acid sequences. In the chronic ischemic heart disease (CIHD), these age-dependent proteins were found in 50% of cases (in the age group 31-40 years), while in the control group such combination was detected only in 10% of the examined individuals. Subsequent studies in this field would probably reveal molecular mechanisms responsible for impairment and/or ageing of the cardiac muscle and also of the adaptation-dysadaptations mechanisms in the CIHD.

[Identification of AGR2 protein, a novel potential cancer marker, using proteomics technologies].

A comparative analysis of the proteins in prostate tissues of the patients operated for hyperplasia (n = 7) or cancer (n = 5) was performed aiming to search for protein diagnostic markers. Differences in several minor proteins were detected using two-dimensional electrophoresis according to O'Farrel; among them, an additional protein with a molecular weight of 19 kDa and an isoelectric point of 9.0 was observed in four of the cancer cases. Mass spectrometry allowed this protein to be identified as the androgen-induced secreted protein AGR2. The possibility of using AGR2 as a diagnostic marker of prostate cancer is discussed.

[New approaches to molecular diagnosis of prostatic cancer].

We compared prostatic proteins in patients operated for adenoma or cancer. 630 protein fractions were obtained from each tissue sample after fractionation by two-dimentional electrophoresis according to O'Farrell. Comparison of the samples from adenoma and cancer showed their difference by 7 proteins among which were isoforms of glyceraldehydes-3-phosphate-dehydrogenase, alpha-collagen and several little known proteins. Most of the cancer patients had the protein with molecular mass 19 kDa and isoelectric point 9.0. By the results of mass-spectrometry this protein was identified as androgen-induced secreted protein AGR2. This protein is considered a potential oncomarker. Prospects of some postgenome technologies for detection of new diagnostic markers of prostatic cancer are discussed.

[Gelatinase activity of protein fractions obtained by electrophoresis in polyacrylamide gel]. / Izuchenie zhelatinaznoi aktivnosti v belkovykh fraktsiiakh, poluchennykh élektroforezom v poliakrilamidnom gele.

The modification of a method of determination gelatinolytic activity in protein fractions obtained by one and two-dimensional polyacrylamide gel electrophoresis is presented. The presence of a fraction 115-125 kD among human proteins of cortex matter and glomerulus of kidney and also in myocardium having gelatinolytic activity is demonstrated.

[Detection of differences in liver protein composition between recombinant strains of mice with different ThioTEPA sensitivities]. / Vyiavlenie razlichii v sostave belkov pecheni u myshei rekombinantnykh linii, otlichaiushchikhsia po chuvstvitel'nosti k vozdeistviiu tioTEF>

The sensitivity of recombinant mouse strains subjected to 23-27 generations of inbreeding to the clastogenic effect of thioTEPA (triethylene thiophosphoramide) was reestimated, and their characteristics were confirmed. Six 1XC3 recombinant strains were obtained from crossing strains 101/H x C3H/Sn, which differed from one another with respect to the sensitivity to thioTEPA. The protein composition of the liver tissue was studied in the recombinant strains by means of two-dimensional electrophoresis. Interstrain differences with respect to five liver proteins were found, which were correlated with the differences in the response to the mutagen.

[Two-dimensional electrophoresis of myocardial proteins in human cardio-vascular diseases]. / Dvumerhyi élektroforeticheskii analiz belkov miokarda pri nekotorykhformakh serdechno-sosudistoi patologii cheloveka.

The protein composition of human myocardium at some cardiovascular pathologies was studied by use of 2D electrophoresis. It was found that dilitation(al) cardiomyopathy and ischemia are both characterized by transferrin accumulation in myocardium and by enhanced expression of a protein with Mm 3 kD/pI 5. Besides that, at ischemic disease there was seen an elevation of the fetal isoform in the light chain of myosin. For one of the polymorphous protein systems, the occurrence rate changes were also recorded.

[DNA-methylase activity in human cardiac muscle. Association of DNA methylase activity with actin protein fractions]. / DNK-metilaznaia aktivnost' v serdechnoi myshtse cheloveka. Assotsiatsiia DNK-metilaznoi aktivnosti s fraktsiei aktinovykh belkov.

The column isoelectrofocusing activity of the nuclear extracts of the human cardiac muscle has revealed at pH 3.5-8.2 5 peaks of DNA-methylase. When one of these peaks (II) was analyzed by the two-dimensional gel electrophoresis 6 proteins (10, 25, 35, 43, 67 and 120 kDa) were separated. 43 kDa protein had electrophoretic properties similar to actins and was able to methylate cytosine in the DNA molecules. The comparative computer analysis of the primary structure of human actins and several bacterial DNA-methylases has shown the homology of the extensive fragments of these molecules.

[Change in the composition of human skeletal muscles in amyotrophic lateral sclerosis]. / Izuchenie belkovogo sostava skeletnoi myshtsy cheloveka pri bokovom amiotroficheskom skleroze.

Protein composition of human skeletal muscle impaired with lateral amyotropic sclerosis was studied by means of two-dimensional electrophoresis. Some protein fractions were altered. Characteristic property of all the preparations studied proved to be disappearance of three protein fractions of 35 kDa molecular mass and with pI 5.9, 6.0 and 6.1.

[Study of human lymphocyte proteins using two-dimensional electrophoresis]. / Izuchenie belkov limfotsitov cheloveka s pomoshch'iu dvumernogo élektroforeza.

Using O'Farrell 2DE modified method, a panel of human lymphocyte proteins have been studied. A 2-D map involving 273 fractions characterized in M(r)/pI coordinates has been developed. Some proteins were identified on the map and some comparability of experimental results to the Celis et al. data demonstrated. Ten protein fractions displaying some variability in a sample of 30 patients were revealed. A change in the amount of the E2 fraction M(r)/pI -51.0/5.70 was found among patients with chromosome 5 short arm deletion and translocation.

[A comparative analysis of the protein composition of human skeletal and cardiac muscle by two-dimensional electrophoresis]. / Sravnitel'nyi analiz belkovogo sostava skeletnoi i serdechnoi myshtscheloveka dvumernym élektroforezom.

Using a modified O'Farrell 2DE technique, the protein composition of human skeletal muscles was compared to that of human myocardium. The variability of protein composition was analysed in nine types of skeletal muscle tissue. A number of skeletal and heart muscles proteins were identified on the electrophoregrams. Both common and tissue-specific groups of muscle tissue polypeptides were revealed.