Enzyme variants in biosynthesis and biological assessment of different molecular weight hyaluronan.

In the present study, low- and high-molecular-weight hyaluronic acids (LMW-HA and HMW-HA) were synthesized in vitro by truncated Streptococcus equisimilis hyaluronan synthases (SeHAS). The enzyme kinetic parameters were determined for each enzyme variant. The MW, structure, dispersity, and biological activity of polymers were determined by electrophoresis, FTIR spectroscopy, carbazole, cell proliferation, and cell migration assay, respectively. The specific activities were calculated as 7.5, 6.8, 4.9, and 2.8 µgHA µgenzyme-1 min-1 for SeHAS, HAS123, HAS23, and HASIntra, respectively. The results revealed SeHAS produced a polydisperse HMW-HA (268 kDa), while HAS123 and HAS23 produced a polydisperse LMW-HA (< 30 kDa). Interestingly, HASIntra produced a low-disperse LMW-HA. Kinetics studies revealed the truncated variants displayed increased Km values for two substrates when compared to the wild-type enzyme. Biological assessments indicated all LMW-HAs showed a dose-dependent proliferation activity on endothelial cells (ECs), whereas HMW-HAs exhibited an inhibitory effect. Also, LMW-HAs had the highest cell migration effect at 10 µg/mL, while at 200 µg/mL, both LMW- and HMW-HAs postponed the healing recovery rate. The study elucidated that the transmembrane domains (TMDs) of SeHAS affect the enzyme kinetics, HA-titer, HA-size, and HA-dispersity. These findings open new insight into the rational engineering of SeHAS to produce size-defined HA.

Evaluation of transmembrane domain deletions on hyaluronic acid polymerization of hyaluronan synthase isolated from Streptococcus equisimilis group G.

The membrane enzyme of hyaluronan synthase (HAS) is the key enzyme in hyaluronic acid (HA) biosynthesis by coupling UDP-sugars. Prior studies proposed the C-terminus region of HAS enzyme mediates the production rate and molecular weight of HA. The current study describes the isolation and characterizations of a transmembrane HAS enzyme isolated from Streptococcus equisimilis Group G (GGS-HAS) in vitro. The effect of transmembrane domains (TMDs) on HA productivity was determined and the shortest active variant was also identified by recombinant expression of full-length and five truncated forms of GGS-HAS in Escherichia coli. We found that the GGS-HAS enzyme is longer than that of S. equisimilis group C (GCS-HAS) which includes three more residues (LER) at the C-terminus region (positions 418-420) and also one-point mutation at position 120 (E120D). Amino acid sequence alignment demonstrated 98% and 71% identity of GGS-HAS with that of S. equisimilis Group C and S. pyogenes Group A, respectively. The in vitro productivity of the full-length enzyme was 35.57 µg/nmol, however, extended TMD deletions led to a reduction in the HA productivity. The HAS-123 variant showed the highest activity among the truncated forms, indicating the essential role of first, second, and third TMDs for the full activity. Despite a decline in activity, the intracellular variant can still mediate the binding and polymerization of HA without any need for TMDs. This significant finding suggests that the intracellular domain is the core for HA biosynthesis in the enzyme and other domains are probably involved in other attributes including the enzyme kinetics that affect the size distribution of the polymer. However, more investigations on the recombinant forms are still needed to confirm clearly the role of each transmembrane domain on these properties.

Introduction of Novel Drug Targets against Staphylococcus aureus and Proposing Putative Inhibitors against Adenine N1 (m1A22)-tRNA Methyltransferase (TrmK) using Computer-aided Drug Discovery.

BACKGROUND: Nowadays, the emergence of methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant S. aureus (VRSA) strains has dramatically restricted the treatment options against this microorganism. AIM: In this study, we aimed to discover new drug targets and inhibitors against S. aureus. METHODS: This study consists of two major sections. In the upstream evaluation, after a comprehensive coreproteome analysis, essential cytoplasmic proteins with no similarity to the human proteome were selected. Then the S. aureus metabolome-specific proteins were selected, and novel drug targets were identified using the DrugBank database. In the downstream analysis, a structure-based virtual screening approach was performed to reveal potential hit compounds against adenine N1 (m(m1A22)-tRNA methyltransferase (TrmK) using the StreptomeDB library and AutoDock Vina software. The compounds with a binding affinity > -9 kcal/mol were analyzed based on ADMET properties. Finally, the hit compounds were selected based on Lipinski's rule of five (RO5). RESULTS: Three proteins, including glycine glycosyltransferase (FemA), TrmK, and heptaprenyl pyrophosphate synthase subunit A (HepS1), were selected as feasible and promising drug targets based on PDB file availability and their essential role in the survival of the S. aureus. Finally, seven hit compounds, including Nocardioazine_ A, Geninthiocin_D, Citreamicin_delta, Quinaldopeptin, Rachelmycin, Di-AFN_A1 and Naphthomycin_ K were introduced against the binding cavity of TrmK, as a feasible drug target. CONCLUSION: The results of this study provided three feasible drug targets against S. aureus. In the following, seven hit compounds were introduced as potential inhibitors of TrmK, and Geninthiocin_D was identified as the most desirable agent. However, in vivo and in vitro investigations are needed to confirm the inhibitory effect of these agents on S. aureus.

Synthesis and characterization of Gd3+-loaded hyaluronic acid-polydopamine nanoparticles as a dual contrast agent for CT and MRI scans.

Magnetic resonance imaging and computed tomography (CT) suffer from low contrast sensitivity and potential toxicity of contrast agents. To overcome these limitations, we developed and tested a new class of dual contrast agents based on polydopamine nanoparticles (PDA-NPs) that are functionalized and targeted with hyaluronic acid (HA). These nanoparticles (NPs) are chelated with Gd3+ to provide suitable contrast. The targeted NPs were characterized through ultraviolet-visible spectroscopy (UV-vis), scanning electron microscopy (SEM), infrared Fourier transform (FTIR), and dynamic light scattering (DLS). The cytotoxicity was investigated on HEK293 cells using an MTT assay. The contrast property of synthesized Gd3+/PDA/HA was compared with Barium sulfate and Dotarem, as commercial contrast agents (CAs) for CT and MRI, respectively. The results illustrated that synthesized PDA-NPs have a spherical morphology and an average diameter of 72 nm. A distinct absorption peak around 280 nm in the UV-vis spectrum reported the self-polymerization of PDA-NPs. The HA coating on PDA-NPs was revealed through a shift in the FTIR peak of C=O from 1618 cm-1 to 1635 cm-1. The Gd3+ adsorption on PDA/HA-NPs was confirmed using an adsorption isotherm assay. The developed CA showed low in vitro toxicity (up to 158.98 µM), and created a similar contrast in MRI and CT when compared to the commercial agents. The r1 value for PDA/HA/Gd3+ (6.5 (mg/ml)-1 s-1) was more than Dotarem (5.6 (mg/ml)-1 s-1) and the results of the hemolysis test showed that at concentrations of 2, 4, 6, and 10 mg/ml, the hemolysis rate of red blood cells is very low. Additionally, the results demonstrated that PDA/HA/Gd3+ could better target the CD44+-expressing cancer cells than PDA/Gd3+. Thus, it can be concluded that lower doses of developed CA are needed to achieve similar contrast of Dotarem, and the developed CA has no safety concerns in terms of hemolysis. The stability of PDA/HA/Gd3+ has also been evaluated by ICP-OES, zeta potential, and DLS during 3 days, and the results suggested that Gd-HA NPs were stable.

Pharmacological effects and therapeutic potential of natural compounds in neuropsychiatric disorders: An update.

Neuropsychiatric diseases are a group of disorders that cause significant morbidity and disability. The symptoms of psychiatric disorders include anxiety, depression, eating disorders, autism spectrum disorders (ASD), attention-deficit/hyperactivity disorder, and conduct disorder. Various medicinal plants are frequently used as therapeutics in traditional medicine in different parts of the world. Nowadays, using medicinal plants as an alternative medication has been considered due to their biological safety. Despite the wide range of medications, many patients are unable to tolerate the side effects and eventually lose their response. By considering the therapeutic advantages of medicinal plants in the case of side effects, patients may prefer to use them instead of chemical drugs. Today, the use of medicinal plants in traditional medicine is diverse and increasing, and these plants are a precious heritage for humanity. Investigation about traditional medicine continues, and several studies have indicated the basic pharmacology and clinical efficacy of herbal medicine. In this article, we discuss five of the most important and common psychiatric illnesses investigated in various studies along with conventional therapies and their pharmacological therapies. For this comprehensive review, data were obtained from electronic databases such as MedLine/PubMed, Science Direct, Web of Science, EMBASE, DynaMed Plus, ScienceDirect, and TRIP database. Preclinical pharmacology studies have confirmed that some bioactive compounds may have beneficial therapeutic effects in some common psychiatric disorders. The mechanisms of action of the analyzed biocompounds are presented in detail. The bioactive compounds analyzed in this review are promising phytochemicals for adjuvant and complementary drug candidates in the pharmacotherapy of neuropsychiatric diseases. Although comparative studies have been carefully reviewed in the preclinical pharmacology field, no clinical studies have been found to confirm the efficacy of herbal medicines compared to FDA-approved medicines for the treatment of mental disorders. Therefore, future clinical studies are needed to accelerate the potential use of natural compounds in the management of these diseases.

Prognostic Value and Biological Role of miR-126 in Breast Cancer.

In eukaryotic organisms such as humans, some noncoding single-stranded RNAs (ncRNAs) contribute to regulating the expression of some genes before and after the transcription process, which in turn controls a number of vital physiological processes, including cell proliferation, differentiation, invasion, angiogenesis, and embryonic development. miR-126 is one of these miRNAs expressed exclusively in endothelial cells such as capillaries and vessels involved in controlling angiogenesis. In recent years, the link between miRs such as miR-126 and the pathology of breast cancer has attracted the attention of many researchers. Numerous studies have shown that miR-126 may be able to suppress tumor tissue metastasis or to increase tumor metastasis through complex molecular mechanisms. There is ample clinical evidence that miR-126 can be used as a biomarker to predict and diagnose breast cancer due to the increased or decreased expression of certain genes in breast cancer tissue. In this review, we discuss the association between the growth and metastasis (tumorigenesis) of breast cancer and miR-126, as well as the relationship between current research advances in the prognosis, diagnosis, and treatment of breast cancer and miR-126.

Withdrawal Notice: Overview of Enzyme Engineering Towards the Production of Hyaluronic Acid with Tailored Molecular Weight

The article has been withdrawn at the request of the author and the editor of the journal Current Protein and Peptide Science.Bentham Science apologizes to the readers of the journal for any inconvenience this may cause.The Bentham Editorial Policy on Article Withdrawal can be found at https://benthamscience.com/editorial-policiesmain.php BENTHAM SCIENCE DISCLAIMER: It is a condition of publication that manuscripts submitted to this journal have not been published and will not be simultaneously submitted or published elsewhere. Furthermore, any data, illustration, structure or table that has been published elsewhere must be reported, and copyright permission for reproduction must be obtained. Plagiarism is strictly forbidden, and by submitting the article for publication the authors agree that the publishers have the legal right to take appropriate action against the authors, if plagiarism or fabricated information is discovered. By submitting a manuscript, the authors agree that the copyright of their article is transferred to the publishers if and when the article is accepted for publication.

Transplantation of Stem Cells as a Potential Therapeutic Strategy in Neurodegenerative Disorders.

Stem cells are considered to have significant capacity to differentiate into various cell types in humans and animals. Unlike specialized cells, these cells can proliferate several times to produce millions of cells. Nowadays, pluripotent stem cells are important candidates to provide a renewable source for the replacement of cells in tissues of interest. The damage to neurons and glial cells in the brain or spinal cord is present in neurological disorders such as Amyotrophic lateral sclerosis, stroke, Parkinson's disease, multiple sclerosis, Alzheimer's disease, Huntington's disease, spinal cord injury, lysosomal storage disorder, epilepsy, and glioblastoma. Therefore, stem cell transplantation can be used as a novel therapeutic approach in cases of brain and spinal cord damage. Recently, researchers have generated neuron-like cells and glial-like cells from embryonic stem cells, mesenchymal stem cells, and neural stem cells. In addition, several experimental studies have been performed for developing stem cell transplantation in brain tissue. Herein, we focus on stem cell therapy to regenerate injured tissue resulting from neurological diseases and then discuss possible differentiation pathways of stem cells to the renewal of neurons.

Epigenetic modifications in gastric cancer: Focus on DNA methylation.

Gastric cancer is a complex heterogeneous disease which is the fourth prevalent malignancy worldwide. Although, several diagnosis and treatment are available for the gastric cancer patients, however the malignancy is still the third leading cause of cancer-related death in the world. Beside the genetic and environmental factors, epigenetic alterations are also involved in the emergence of gastric cancer. Epigenetics alterations are heritable changes which regulate gene expression without occurring changes in DNA sequence. Epigenetic changes mostly include DNA methylation, histon post-translational modifications, chromatin remodeling and non-coding RNAs. Among the mentioned epigenetic modifications, DNA methylation is a major epigenetic process that plays a key role in different stages of evolution and cancer development. In this review, we address all types of related epigenetic modifications in gastric cancer by focus on DNA methylation by reviewing the recent literatures. Understanding of molecular mechanisms of epigenetics alterations in gastric cancer development helps researchers to identify new epigenetic drugs against the malignancy.

High doses of garlic extract significantly attenuated the ratio of serum LDL to HDL level in rat-fed with hypercholesterolemia diet.

BACKGROUND: Hypercholesterolemia is associated with an increased risk of heart disease. In this study, we investigated the antihyperlipidemic effects of garlic (Allium sativum L.) in rat models of hypercholesterolemic. METHODS: Wistar male rats were randomly divided into 4 diet groups with garlic supplementation. Male Wistar rats were fed by standard pellet diet (group I), standard diet supplemented with 4% garlic (group II), lipogenic diet (containing sunflower oil, cholesterol and ethanol) equivalent to 200 mg raw garlic/kg body weight (raw) (group III) and lipogenic diet equivalent to 400 mg raw garlic/kg body weight (raw) (group IV). RESULTS: Rats fed 400 g/kg garlic extract(GE), had a significantly lower concentration of serum low-density lipoprotein cholesterol (LDL-C) cholesterol and elevated HDL -C cholesterol at day 28 (P < 0.05).In addition,serum levels of LDL-C was lower in the III and IV group than those in the IV group (P < 0.001 for each). However, cholesterol efflux capacity was positively correlated with HDL cholesterol concentration (P < 0 · 0001). It was also directly correlated with garlic supplementation (P < 0 · 0001). CONCLUSION: Together Taken, the results are clearly indicative of the beneficial effects of garlic in reducing lateral side effects of hyperlipidemia. Our data demonstrate that GE has protective effects on HDL in rats with high LDL intake. Therefore, it could be used to remedy hypercholesterolemia with help reduce risk of coronary heart disease VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1834155749171141.

Comparison of the metal concentrations in the feathers of three bird species from southern Iran.

This study was conducted to determine the concentration of metals, namely cadmium, lead, cobalt and copper, in the feathers of chukar (Alectoris chukar), see-see partridge (Ammoperdix griseogularis) and rock dove (Columba livia) in order to: examine the species, gender and age related variations in trace metal accumulation, and identify any relationships between species. Bird samples were collected in February 2012 from the Hormod protected area, southern Iran and the concentration of metals were measured using a Shimadzu AA 660 flame atomic absorption spectrophotometer. The concentrations in all three species were copper > lead > cadmium > cobalt. The average cadmium concentrations were 2.0, 1.9, and 1.9 µg/g for A. chukar, A. griseogularis, and C. livia, respectively. The average lead concentrations were 8.0, 5.4, and 7.7 µg/g for A. chukar, A. griseogularis, and C. livia, respectively. The results showed that in all three species, the highest metal concentrations were observed in female/adult birds and the lowest concentrations were found in female/juvenile birds (except lead in A. chukar and copper in C. livia).