Effect of genetic variants of chemokine receptors on the development of myocardial infarction in Turkish population

Inflammation is a crucial component of coronary atherosclerosis and myocardial infarction (MI). Chemokine receptors are important modulators of inflammation. Polymorphisms in genes coding for chemokine receptors, CCR2 and CCR5, have been studied as genetic markers of coronary artery disease. In the present study, we investigated whether genetic variants of CCR2-V64I and CCR5-delta32 chemokine receptors have any effect on the development of myocardial infarction. A total of 146 MI patients and 202 control subjects were genotyped for CCR2 and CCR5. CCR2-V64I genotypes were not significantly different between patients with MI and controls (P > 0.05). CCR5-delta32 genotype distribution in cases was significantly different from that of controls (P = 0.042). The CCR5-delta32 wt/deletion genotype frequencies for controls and cases were 0.10 and 0.19, respectively and individuals with CCR5-delta32 wt/deletion genotype had a 2.13-fold increased risk of myocardial infarction (P = 0.0013). Individuals carrying the CCR5-delta32 heterozygote or homozygous variant genotype (deletion/deletion + wt/deletion) had a 1.96-fold increased risk of myocardial infarction compared with the wild-type genotype (wt/wt) (p: 0.016). In conclusion, our data have suggested that genetic variant of CCR5 might be associated with the development of MI. Further larger sample size studies are required to confirm our findings.     

Purification and Characterisation of Rat Kidney Glutathione Reductase

Glutathione reductase [GR, E.C.1.8.1.7] catalyses NADPH dependent reduction of glutathione disulfide (GSSG) to reduced glutathione (GSH). Thus, it is the crucial enzyme to maintain high [GSH]/[GSSG] ratio and physiological redox status in cells. Kidney and liver tissues were considered as a rich source of GR. In this study, rat kidney GR was purified and some of its properties were investigated. The enzyme was purified 2,356 fold with a yield of 16% by using heat-denaturation and Sephadex G25 gel filtration, 2′,5′-ADP Agarose 4B, PBE94 column chromatographies. The purified enzyme had a specific activity (Vm) of 250 U/mg protein and the ratio of absorbances at wavelengths of A ₂₇₃/A _463, A ₂₈₀/A ₄₆₀, A ₃₆₅/A ₄₆₀, and A ₃₇₉/A ₄₆₃, were 7.1, 6.8, 1.2 and 1.0, respectively. Each mol of GR subunit bound 0.97 mol of FAD. NADH was used as a coenzyme by rat kidney GR but with a lower efficiency (32.7%) than NADPH. Its subunit molecular weight was estimated as 53 kDa. An optimum pH of 6.5 and optimum temperature of 65 °C were found for rat kidney GR. Its activation energy (Ea) and temperature coefficient (Q₁₀) were calculated as 7.02 kcal/mol and 1.42, respectively. The Km_(NADPH) and kcat/Km _(NADPH) values were found to be 15.3 ± 1.4 μM and 1.68 × 10⁷ M⁻¹ s⁻¹ for the concentration range of 10-200 μM NADPH and when GSSG is the variable substrate, the Km_(GSSG) and the kcat/Km_(GSSG) values of 53.1 ± 3.4 μM and 4.85 × 10⁶ M⁻¹ s⁻¹ were calculated for the concentration range of 20–1,200 μM GSSG.     

Evaluation of in vitro Probiotic Potential of Pediococcus pentosaceus OZF Isolated from Human Breast Milk

This study was conducted to evaluate the probiotic properties of Pediococcus pentosaceus OZF isolated from human breast milk. The results obtained so far suggest that the strain is resistant to low pH, bile salt, pepsin and pancreatin, so it could survive while passing through the upper part of the gastrointestinal tract and reveal its potential probiotic action on host organism. The strain was non-pathogenic (γ-hemolytic), produced anti-Listerial bacteriocin, exhibited a strong autoaggregating phenotype (85.71%) and demonstrated 6.26 and 12.99% coaggregation with Salmonella enterica serotype Typhimurium SL 1344 and Escherichia coli LMG 3083 (ETEC), respectively. The degree of adhesion of Ped. pentosaceus OZF to the human Caco-2 cell line was investigated and when compared to the adhesion of pathogenic strains tested, it was shown to inhibit the growth of human enterotoxigenic E. coli LMG 3083 (ETEC) and of Salm. Typhimurium SL 1344. Ped. pentosaceus OZF seems to adhere to human intestinal cells via mechanisms that involve different combinations of carbohydrate and lipid factors on the bacteria and eukaryotic cell surface. The percentage of adhesion to n-hexadecane was 34% showing that the surface was rather hydrophilic. Higher affinity displayed by Ped. pentosaceus OZF for chloroform demonstrates the basic property of a cell, which may be due to the presence of carboxylic groups on the cell surface.     

Kinetic properties and storage stability of catalase immobilized on to florisil

The covalent immobilization of bovine liver catalase (CAT) on to florisil via glutaraldehyde was investigated. Optimum immobilization pH and temperature were determined as pH 6.0, 10 degrees C respectively, while the amount of initial CAT per g of carrier and immobilization time was determined as 5 mg g(-1) and 120 min, respectively. The Vmax values for free and immobilized CAT were found to be 1.7 x 10(5) and 2.0 x 10(4) micromol H2O2 min(-1) mg protein(-1), respectively, whereas KM values were 33.3 mM and 1722.0 mM respectively. Operational stability was determined by using a stirred batch-type column reactor. Immobilized CAT retained about 40% of its initial activity after 50 uses. It showed higher storage stability than free CAT at 4 degrees C and 25 degrees C. Its storage stability increased with increasing relative humidity (RH) from 0 to 20% of the medium. The highest storage stability was obtained in 20% RH, however, further increase in RH from 40 to 100% significantly decreased the storage stability.     

Paraoxonase and arylesterase activities in fibromyalgia

We aimed to evaluate the association of serum paraoxonase and arylesterase activities and oxidative/antioxidative status in patients with fibromyalgia. Forty-two patients with fibromyalgia and 53 healthy controls were included in the study. Serum paraoxonase and arylesterase activities were measured spectrophotometrically. Oxidative and antioxidative status were evaluated by measuring serum lipid hydroperoxide (LOOH) levels, total antioxidant status (TAS) and free sulfhydryl groups (-SH = total thiol). Lipid parameters were determined by routine laboratory methods. Serum paraoxonase and arylesterase activities, and TAS were lower in patients with fibromyalgia than in controls (P < 0.001, for all), and the -SH level was also lower in the patient group (P = 0.03). LOOH levels were higher in the patient group than in controls (P = 0.01). Our results suggest that patients with fibromyalgia were exposed to oxidative stress, and paraoxonase and arylesterase activities were decreased in these patients. Patients with fibromyalgia might be prone to development of atherosclerosis with reduced paraoxonase and arylesterase activities.     

Structural cooperativity in histone H3 tail modifications

Post-translational modifications of histone H3 tails have crucial roles in regulation of cellular processes. There is cross-regulation between the modifications of K4, K9, and K14 residues. The modifications on these residues drastically promote or inhibit each other. In this work, we studied the structural changes of the histone H3 tail originating from the three most important modifications; tri-methylation of K4 and K9, and acetylation of K14. We performed extensive molecular dynamics simulations of four types of H3 tails: (i) the unmodified H3 tail having no chemical modification on the residues, (ii) the tri-methylated lysine 4 and lysine 9 H3 tail (K4me3K9me3), (iii) the tri-methylated lysine 4 and acetylated lysine 14 H3 tail (K4me3K14ace), and (iv) tri-methylated lysine 9 and acetylated lysine 14 H3 tail (K9me3K14ace). Here, we report the effects of K4, K9, and K14 modifications on the backbone torsion angles and relate these changes to the recognition and binding of histone modifying enzymes. According to the Ramachandran plot analysis; (i) the dihedral angles of K4 residue are significantly affected by the addition of three methyl groups on this residue regardless of the second modification, (ii) the dihedral angle values of K9 residue are similarly altered majorly by the tri-methylation of K4 residue, (iii) different combinations of modifications (tri-methylation of K4 and K9, and acetylation of K14) have different influences on phi and psi values of K14 residue. Finally, we discuss the consequences of these results on the binding modes and specificity of the histone modifying enzymes such as DIM-5, GCN5, and JMJD2A.     

A comparative molecular dynamics study of methylation state specificity of JMJD2A

Histone modifications have great importance in epigenetic regulation. JMJD2A is a histone demethylase which is selective for di- and trimethyl forms of residues Lys9 and Lys36 of Histone 3 tail (H3K9 and H3K36). We present a molecular dynamics simulations of mono-, di- and trimethylated histone tails in complex with JMJD2A catalytic domain to gain insight into how JMJD2A discriminates between the methylation states of H3K9. The methyl groups are located at specific distances and orientations with respect to Fe(II) in methylammonium binding pocket. For the trimethyllysine the mechanism which provides the effectual orientation of methyl groups is the symmetry, whereas for the dimethyllysine case the determining factors are the interactions between methyllysine head and its environment and subsequently the restriction on angular motion. The occurrence frequency of methyl groups in a certain proximity of Fe(II) comes out as the explanation of the enzyme activity difference on di- and tri-methylated peptides. Energy analysis suggests that recognition is mostly driven by van der Waals and followed by Coulombic interactions in the enzyme-substrate interface. The number (mono, di or tri) and orientations of methyl groups and water molecules significantly affect the extent of van der Waals interaction strengths. Hydrogen bonding analysis suggests that the interaction between JMJD2A and its substrates mainly comes from main chain-side chain interactions. Binding free energy analysis points out Arg8 as an important residue forming an intra-substrate hydrogen bond with tri and dimethylated Lys9 of the H3 chain. Our study provides new insights into how JMJD2A discriminates between its substrates from both a structural and dynamical point of view.