The effect of caffeic acid phenethyl ester (CAPE) on metabolic enzymes including acetylcholinesterase,butyrylcholinesterase, glutathione S-transferase,lactoperoxidase, and carbonic anhydrase isoenzymes I,II, IX,and XII

Caffeic acid phenethyl ester (CAPE) is an active component of honeybee propolis extracts. Carbonic anhydrases (CAs, EC 4.2.1.1) are widespread and intensively studied metalloenzymes present in higher vertebrates including humans as many diverse isoforms. Acetylcholinesterase (AChE) is responsible for acetyl choline (ACh) hydrolysis and plays a fundamental role in nerve impulse transmission by terminating the action of the ACh neurotransmitter at cholinergic synapses and neuromuscular junctions. Butyrylcholinesterase (BChE) is another enzyme abundantly present in the liver and released into blood in a soluble form. Lactoperoxidase (LPO) is an enzyme involved in fighting pathogenic microorganisms whereas glutathione S-transferases (GSTs) are dimeric proteins present both in prokaryotic and eukaryotic organisms and involved in cellular detoxification mechanisms. In the present study, the inhibition effect of CAPE on human carbonic anhydrase (hCA) isoforms I, II, IX, and XII, AChE, BChE, LPO, and GST was evaluated. CAPE inhibited these enzymes with K_is in the range between micromolar to picomolar. The best inhibitory effect was observed against AChE and BChE.     

Enantioselective Ethylation of Various Aldehydes Catalyzed by Readily Accessible Chiral Diols

Four chiral C₂‐symmetric diols were synthesized in a straightforward three‐step reaction and demonstrated excellent enantioselectivities and good overall yields. Their catalytic activities were examined via the addition of diethylzinc to various aldehydes. The enantioselective addition of diethylzinc to 2‐methoxybenzaldehyde gave the corresponding chiral secondary alcohol with high yields (up to 95%) and moderate to good enantiomeric excess (up to 88%). All synthesized ligands were evaluated in the addition of diethylzinc to various aldehydes in the presence of an additional metal such as Ti(IV) complexes. Chirality 28:593–598, 2016. © 2016 Wiley Periodicals, Inc.     

Detection and Identification of Phytoplasmas in Pomegranate Trees with Yellows Symptoms

Symptoms resembling those associated with phytoplasma presence were observed in pomegranate (Punica granatum L.) trees in June 2012 in the Aegean Region of Turkey (Ayd?n province). The trees exhibiting yellowing, reduced vigour, deformations and reddening of the leaves and die‐back symptoms were analysed to verify phytoplasma presence. Total nucleic acids were extracted from fresh leaf midribs and phloem tissue from young branches of ten symptomatic and five asymptomatic plants. Nested polymerase chain reaction assays using universal phytoplasma‐specific 16S rRNA and tuf gene primers were performed. Amplicons were digested with Tru1I, Tsp509I and HhaI restriction enzymes, according to the primer pair employed. The phytoplasma profiles were identical to each other and to aster yellows (16SrI‐B) strain when digestion was carried out on 16Sr(I)F1/R1 amplicons. However, one of the samples showed mixed profiles indicating that 16SrI‐B and 16SrXII‐A phytoplasmas were present when M1/M2 amplicons were digested, the reamplification of this sample with tuf cocktail primers allowed to verify the presence of a 16SrXII‐A profile. One pomegranate aster yellows strain AY‐PG from 16S rRNA gene and the 16SrXII‐A amplicon from tuf gene designed strain STOL‐PG were directly sequenced and deposited in GenBank under the Accession Numbers KJ818293 and KP161063, respectively. To our knowledge, this is the first report of 16SrI‐B and 16SrXII‐A phytoplasmas in pomegranate trees.     

Tailoring of recombinant FDH: effect of histidine tag location on solubility and catalytic properties of Chaetomium thermophilum formate dehydrogenase (CtFDH)

Abstract

Several protein expression systems can be used to get enzymes in required quantities and study their functions. Incorporating a polyhistidine tag is a beneficial way of getting various enzymes such as FDHs for industrial applications. The NAD⁺ dependent formate dehydrogenase from Chaetomium thermophilum (CtFDH) can be utilized for interconversion of formate to carbon dioxide coupled with the conversion of NAD⁺ to NADH. In this study, N-terminal His tagged CtFDH (N-CtFDH) and C-terminal His tagged CtFDH (C-CtFDH) was constructed to learn the effect of His tag location on the activity and kinetic parameters of the enzyme. The solubility of proteins was not affected by tag position, however, an interference on the N-terminal region caused a deterioration in specific activity and the kinetic ability of enzyme. The obtained results indicated that the C-terminus of the enzyme is an appropriate region for tag engineering. The C-CtFDH has an approximately three-fold larger specific activity and two-fold higher catalytic efficiency than N-CtFDH. The results suggest that insertion of a His-tag at the N-terminal or C-terminal end of CtFDH has different effects on the protein and the N-terminal fragment of the protein is crucial for the function of CtFDH.     

Synthesis and utilization of reversible and irreversible light-activated nanovalves derived from the channel protein MscL

This protocol details the chemical modification of the mechanosensitive channel of large-conductance (MscL) channel protein into a light-activated nanovalve and its utilization for triggered delivery in synthetic liposomal vesicles. It is based on charge-induced activation of this otherwise mechanosensitive channel by covalent attachment to the protein of rationally designed synthetic functionalities. In the dark, these functionalities will be uncharged and the channel will stay closed, but UV illumination will cause their ionization and trigger channel activity. In the case of reversible activation, subsequent illumination with visible light will neutralize the charge, causing the channel to close. The protocol includes synthesis of light-responsive compounds, protein isolation and its chemical labeling, reconstitution of the protein into artificial membranes, its analysis at the single-molecule level and its application in liposomal delivery. The whole protocol takes 4 days. Unlike mutagenesis, this method allows the introduction of custom-designed functional groups.     

Effects of Quercetin against Doxorubicin-Induced Testicular Toxicity in Male Rats

Biology Bulletin - This study investigated the protective effects of quercetin (QUE) in doxorubicin (DOX)-induced testicular toxicity in rats. In the present study, testicular histomorphometry, the...     

Autophagy and mTOR pathways in mouse embryonic stem cell,lung cancer and somatic fibroblast cell lines

Embryonic developmental stages and regulations have always been one of the most intriguing aspects of science. Since the cancer stem cell discovery, striking for cancer development and recurrence, embryonic stem cells and control mechanisms, as well as cancer cells and cancer stem cell control mechanisms become important research materials. It is necessary to reveal the similarities and differences between somatic and cancer cells which are formed of embryonic stem cells divisions and determinations. For this purpose, mouse embryonic stem cells (mESCs), mouse skin fibroblast cells (MSFs) and mouse lung squamous cancer cells (SqLCCs) were grown in vitro and the differences between these three cell lines signalling regulations of mechanistic target of rapamycin (mTOR) and autophagic pathways were demonstrated by immunofluorescence and real-time polymerase chain reaction. Expressional differences were clearly shown between embryonic, cancer and somatic cells that mESCs displayed higher expressional level of Atg10, Hdac1 and Cln3 which are related with autophagic regulation and Hsp4, Prkca, Rhoa and ribosomal S6 genes related with mTOR activity. LC3 and mTOR protein levels were lower in mESCs than MSFs. Thus, the mechanisms of embryonic stem cell regulation results in the formation of somatic tissues whereas that these cells may be the causative agents of cancer in any deterioration.