Preparation and properties of biologically active radioiodinated parathyroid hormone

A constant-current microelectrolytic radioiodination method was used to label bovine parathyroid hormone (BPTH) with ¹²⁵I to an overall iodination ratio of 1:1 iodide atoms per PTH molecule. Such iodinated preparations were shown to be fully active in several bioassay systems: in vitro adenylate cyclase activation in rat renal and skeletal membranes, in vitro calcium release from rat calvaria, and the in vivo hypercalcemic response in chickens. Analysis by Sephadex G-15 chromatography after enzymatic digestion showed the radioiodine to be incorporated predominantly as monoiodotyrosine. Bioassay of iodinated preparations from which uniodinated hormone had been removed by isoelectric focusing showed essentially full hormonal activity. Such methods can be used to consistently produce radioiodinated biologically active preparations of BPTH 1–84 with high specific activity (2000 Ci/mmol).     

Clinical Implications of MiR128, Angiotensin I Converting Enzyme and Vascular Endothelial Growth Factor Gene Abnormalities and Their Association with T2D

Type 2 DM (T2D) results from the interaction of the genetic and environmental risk factors. Vascular endothelial growth factor (VEGF), angiotensin I-converting enzyme (ACE), and MicroRNAs (MiRNAs) are involved in important physiological processes. Gene variations in VEGF, ACE and MiRNA genes are associated with diseases. In this study we investigated the associations of the VEGF-2578 C/A (rs699947), VEGF-2549 insertion/deletion (I/D), and ACE I/D rs4646994 and Mir128a (rs11888095) gene variations with T2D using the amplification refractory mutation system PCR (ARMS-PCR) and mutation specific PCR (MSP). We screened 122 T2D cases and 126 healthy controls (HCs) for the rs699947, and 133 T2D cases and 133 HCs for the VEGF I/D polymorphism. For the ACE I/D we screened 152 cases and 150 HCs, and we screened 129 cases and 112 HCs for the Mir128a (rs11888095). The results showed that the CA genotype of the VEGF rs699947 and D allele of the VEGF I/D polymorphisms were associated with T2D with OR =2.01, p-value = 0.011, and OR = 2.42, p-value = 0.010, respectively. The result indicated the D allele of the ACE ID was protective against T2D with OR = 0.10, p-value = 0.0001, whereas the TC genotype and the T allele of the Mir128a (rs11888095) were associated with increased risk to T2D with OR = 3.16, p-value = 0.0001, and OR = 1.68, p-value = 0.01, respectively. We conclude that the VEGF (rs699947), VEGF I/D and Mir128a (rs11888095) are potential risk loci for T2D, and that the D allele of the ACE ID polymorphism may be protective against T2D. These results help in identification and stratification for the individuals that at risk for T2D. However, future well-designed studies in different populations and with larger sample sizes are required. Moreover, studies to examine the effects of these polymorphisms on VEGF and ACE proteins are recommended.     

Identification and SAR of novel diaminopyrimidines. Part 1: The discovery of RO-4, a dual P2X3/P2X2/3 antagonist for the treatment of pain

P2X purinoceptors are ligand-gated ion channels whose endogenous ligand is ATP. Both the P2X₃ and P2X_2/3 receptor subtypes have been shown to play an important role in the regulation of sensory function and dual P2X₃/P2X_2/3 antagonists offer significant potential for the treatment of pain. A high-throughput screen of the Roche compound collection resulted in the identification of a novel series of diaminopyrimidines; subsequent optimization resulted in the discovery of RO-4, a potent, selective and drug-like dual P2X₃/P2X_2/3 antagonist.     

The effects of the flood cycle on the diversity and composition of the phytoplankton community of a seasonally flooded Ramsar wetland in Bangladesh

Freshwater wetlands in Bangladesh are strongly influenced by the monsoons and the annual flood cycle has measurable impacts on the abiotic and biotic components of these ecosystems. The northeastern Haor Basin of Bangladesh is particularly rich in seasonally flooded freshwater wetlands that support a wide diversity of flora and fauna. These wetlands are of great importance to the local economy due to the abundance of rich floodplain fisheries. Little is known about the phytoplankton communities of these wetlands that are known to be linked with zooplankton and fish productivity. We investigated the seasonal variation in the diversity and abundance of phytoplankton assemblages in Tanguar Haor, a Ramsar wetland in northeastern Bangladesh during the period of inundation (June–December). A total of 107 genera of phytoplankton representing five classes were recorded. Blooms of Microcystis dominated the phytoplankton community throughout the study period but were particularly acute during the early part of the high water period. Among the Bacillariophyceae, Melosira was the most dominant, reaching bloom proportions early in the high water period. Factor analysis of physicochemical variables separated the flood cycle into four distinct periods: early high water, mid high water, late high water and low water periods. Phase of the flood cycle, nutrient availability, the physicochemical variables combined with the dominance of Microcystis seemed to be important in controlling the abundance, diversity and dynamics of the phytoplankton genera. The abundance of genera of desmids and some Bacillariophyceae is indicative of the relatively unpolluted conditions of Tanguar Haor.     

Spectroscopic and molecular modelling analysis of the interaction between ethane‐1,2‐diyl bis(N,N‐dimethyl‐N‐hexadecylammoniumacetoxy)dichloride and bovine serum albumin

Several spectroscopic approaches namely fluorescence, time‐resolved fluorescence, UV‐visible, and Fourier transform infra‐red (FT‐IR) spectroscopy were employed to examine the interaction between ethane‐1,2‐diyl bis(N,N‐dimethyl‐N‐hexadecylammoniumacetoxy)dichloride (16‐E2‐16) and bovine serum albumin (BSA). Fluorescence studies revealed that 16‐E2‐16 quenched the BSA fluorescence through a static quenching mechanism, which was further confirmed by UV–visible and time‐resolved fluorescence spectroscopy. In addition, the binding constant and the number of binding sites were also calculated. The thermodynamic parameters at different temperatures (298 K, 303 K, 308 K and 313 K) indicated that 16‐E2‐16 binding to BSA is entropy driven and that the major driving forces are electrostatic interactions. Decrease of the α‐helix from 53.90 to 46.20% with an increase in random structure from 22.56 to 30.61% were also observed by FT‐IR. Furthermore, the molecular docking results revealed that 16‐E2‐16 binds predominantly by electrostatic and hydrophobic forces to some residues in the BSA sub‐domains IIA and IIIA. Copyright © 2015 John Wiley & Sons, Ltd.     

Chloroplast localization of Cry1Ac and Cry2A protein- an alternative way of insect control in cotton

Background

Insects have developed resistance against Bt-transgenic plants. A multi-barrier defense system to weaken their resistance development is now necessary. One such approach is to use fusion protein genes to increase resistance in plants by introducing more Bt genes in combination. The locating the target protein at the point of insect attack will be more effective. It will not mean that the non-green parts of the plants are free of toxic proteins, but it will inflict more damage on the insects because they are at maximum activity in the green parts of plants.

Results

Successful cloning was achieved by the amplification of Cry2A, Cry1Ac, and a transit peptide. The appropriate polymerase chain reaction amplification and digested products confirmed that Cry1Ac and Cry2A were successfully cloned in the correct orientation. The appearance of a blue color in sections of infiltrated leaves after 72 hours confirmed the successful expression of the construct in the plant expression system. The overall transformation efficiency was calculated to be 0.7%. The amplification of Cry1Ac-Cry2A and Tp2 showed the successful integration of target genes into the genome of cotton plants. A maximum of 0.673 μg/g tissue of Cry1Ac and 0.568 μg/g tissue of Cry2A was observed in transgenic plants. We obtained 100% mortality in the target insect after 72 hours of feeding the 2^nd instar larvae with transgenic plants. The appearance of a yellow color in transgenic cross sections, while absent in the control, through phase contrast microscopy indicated chloroplast localization of the target protein.

Conclusion

Locating the target protein at the point of insect attack increases insect mortality when compared with that of other transgenic plants. The results of this study will also be of great value from a biosafety point of view.     

Molecular investigation of the interaction between ionic liquid type gemini surfactant and lysozyme: A spectroscopic and computational approach

Herein, we are reporting the interaction of ionic liquid type gemini surfactant, 1,4‐bis(3‐dodecylimidazolium‐1‐yl) butane bromide ([C₁₂?4‐C₁₂im]Br₂) with lysozyme by using Steady state fluorescence, UV‐visible, Time resolved fluorescence, Fourier transform‐infrared (FT‐IR) spectroscopy techniques in combination with molecular modeling and docking method. The steady state fluorescence spectra suggested that the fluorescence of lysozyme was quenched by [C₁₂?4‐C₁₂im]Br₂ through static quenching mechanism as confirmed by time resolved fluorescence spectroscopy. The binding constant for lysozyme‐[C₁₂?4‐C₁₂im]Br₂ interaction have been measured by UV‐visible spectroscopy and found to be 2.541 × 10⁵M^?1. The FT‐IR results show conformational changes in the secondary structure of lysozyme by the addition of [C₁₂?4‐C₁₂im]Br₂. Moreover, the molecular docking study suggested that hydrogen bonding and hydrophobic interactions play a key role in the protein‐surfactant binding. Additionally, the molecular dynamic simulation results revealed that the lysozyme‐[C₁₂?4‐C₁₂im]Br₂ complex reaches an equilibrium state at around 3 ns. © 2015 Wiley Periodicals, Inc. Biopolymers 103: 406–415, 2015.