Glucose Inhibition of the Transport and Phosphoenolpyruvate-Dependent Phosphorylation of Galactose and Fructose in Vibrio cholerae

Vibrio cholerae followed a two-step pattern of growth in a medium containing glucose and either galactose or fructose. Glucose was utilized first. Glucose inhibited the uptake and phosphenolpyruvate-dependent phosphorylation of galactose and fructose.     

Developmentally regulated troponin C mRNAs of chicken skeletal muscle.

Fast and slow/cardiac troponin C (TnC) are the two different isoforms of TnC. Expression of these isoforms is developmentally regulated in vertebrate skeletal muscle. Therefore, in our studies, the pattern of their expression was analyzed by determining the steady-state levels of both TnC mRNAs. It was also examined if mRNAs for both isoforms of TnC were efficiently translated during chicken skeletal muscle development. We have used different methods to determine the steady-state levels of TnC mRNAs. First, probes specific for the fast and slow TnC mRNAs were developed using a 390 base pair (bp) and a 255 bp long fragment, of the full-length chicken fast and slow TnC cDNA clones, respectively. Our analyses using RNA-blot technique showed that fast TnC mRNA was the predominant isoform in embryonic chicken skeletal muscle. Following hatching, a significant amount of slow TnC mRNA began to accumulate in the skeletal (pectoralis) muscle. At 43 weeks posthatching, the slow TnC mRNA was nearly as abundant as the fast isoform. Furthermore, a majority of both slow and fast TnC mRNAs was found to be translationally active. A second method allowed a more reliable measure of the relative abundance of slow and fast TnC mRNAs in chicken skeletal muscle. We used a common highly conserved 18-nucleotide-long sequence towards the 5'-end of these mRNAs to perform primer extension analysis of both mRNAs in a single reaction. The result of these analyses confirmed the predominance of fast TnC mRNA in the embryonic skeletal muscle, while significant accumulation of slow TnC mRNA was observed in chicken breast (pectoralis) muscle following hatching. In addition to primer extension analysis, polymerase chain reaction was used to amplify the fast and slow TnC mRNAs from cardiac and skeletal muscle. Analysis of the amplified products demonstrated the presence of significant amounts of slow TnC mRNA in the adult skeletal muscle.     

Insights from the draft genome of Paenibacillus lentimorbus NRRL B-30488, a promising plant growth promoting bacterium

Paenibacillus lentimorbus NRRL B-30488, a plant growth-promoting bacterium was isolated from Sahiwal cow's milk. The strain shows antagonism against phytopathogens, Fusarium oxysporum f. sp. ciceri and Alternaria solani. Its genome contains gene clusters involved in nonribosomal synthesis of secondary metabolites involved in antimicrobial activities. The genome sequence of P. lentimorbus NRRL B-30488 provides the genetic basis for application of this bacterial strain in plant growth promotion, plant protection and degradation of organic pollutants.     

Vascular imaging in small rodents using micro-CT

In vivo animal models of neoplasm, stroke, subarachnoid hemorrhage, and other diseases involving alterations in vessel anatomy and diameter, require a fast and easy-to-use imaging tool that captures anatomical structure and biologic function data. Micro-computed tomography angiography (μCTA) offers high spatial and temporal resolution and is suitable to perform this task. However, conducting μCTA in small rodents, especially in mice, requires a high degree of accuracy and precision. This article describes a setup for in vivo μCTA in mice using both a bolus technique with a conventional contrast agent, as well as, angiography with a blood-pool contrast agent. Our setup in mice is at isotropic resolutions up to 16 μm with scanning times less than 1 min. The described protocol also addresses some of the technical challenges associated with the imaging of vascular structures in mice models.     

Purification and characterization of a new alkali-thermostable lipase from Staphylococcus aureus isolated from Arachis hypogaea rhizosphere

An extracellular lipase (EC 3.1.1.3), SAL-PP1, from Staphylococcus aureus isolated from Arachis hypogaea rhizosphere was purified and characterized. The enzyme was purified using PALL'S Microsep centrifugal device (10 kD cut off), hydrophobic interaction (phenyl sepharose CL-4B column) and Superose-12 gel filtration chromatography and found to have a molecular mass of around 49 kDa. The gene fragment encoding the part of the catalytic site of the SAL-PP1 lipase was sequenced and the deduced amino acid sequence shows 93% identity with that of SEL3. SAL-PP1 showed activity against long acyl-chain triglycerides, various p-nitrophenyl esters and phospholipids. The enzyme shows high stability and activity after incubation with various metal ions (retained >90% activity in presence of Ca²⁺, Na⁺, Cu²⁺, Mg²⁺, Fe²⁺, or Hg²⁺ at 10 mM), organic solvents (retained >80% activity in presence of acetonitrile, ethanol, DMSO, methanol, isopropanol, toluene, or ethylene glycol at 10 mM), detergents (retained >70% activity in Triton X-100, Tween 80, or sodium deoxycholate at 10 mM) and irreversible inhibitors (retained >77% activity in presence of PMSF, leupetin, or β-mercaptoethanol, at 1 mM). Thermal inactivation studies revealed a temperature dependent unfolding of secondary structure of protein. SAL-PP1 showed maximal activity and stability at pH 8.0 and pH 9.0, respectively. The alkali-thermostability, organic solvent-tolerance and broad substrate specificity of this enzyme may have potential implications in detergent formulations, biotransformation, industries, and medicine.     

Hydrophobic tail length plays a pivotal role in amyloid beta (25–35) fibril–surfactant interactions

The amyloid β‐peptide fragment comprising residues 25–35 (Aβ_25‐35) is known to be the most toxic fragment of the full length Aβ peptide which undergoes fibrillation very rapidly. In the present work, we have investigated the effects of the micellar environment (cationic, anionic, and nonionic) on preformed Aβ_25‐35 fibrils. The amyloid fibrils have been prepared and characterized by several biophysical and microscopic techniques. Effects of cationic dodecyl trimethyl ammonium bromide (DTAB), cetyl trimethylammonium bromide (CTAB), anionic sodium dodecyl sulfate (SDS), and nonionic polyoxyethyleneoctyl phenyl ether (Triton X‐100 or TX) on fibrils have been studied by Thioflavin T fluorescence, UV–vis spectroscopy based turbidity assay and microscopic analyses. Interestingly, DTAB and SDS micelles were observed to disintegrate prepared fibrils to some extent irrespective of their charges. CTAB micelles were found to break down the fibrillar assembly to a greater extent. On the other hand, the nonionic surfactant TX was found to trigger the fibrillation process. The presence of a longer hydrophobic tail in case of CTAB is assumed to be a reason for its higher fibril disaggregating efficacy, the premise of their formation being largely attributed to hydrophobic interactions. Proteins 2016; 84:1213–1223. © 2016 Wiley Periodicals, Inc.     

Enzymatic Substrate Inhibition in Metal Free Catecholase Activity

The catecholase activities were routinely modeled using transition metal complexes as catalyst and in some case basic pH were used as a reaction condition. In this article, the catalytic aerobic oxidation of proxy substrate 3,5-di-tert-butylcatechol (DTBC) in methanol using triethylamine/diethylamine as catalyst was demonstrated as a functional mimic of catecholase activity. The kinetic manifestation of DTBC oxidation was explained as enzymatic substrate inhibition pattern in Michaelis-Menten kinetic model. The mechanistic insight of the aerobic oxidation of DTBC was further validated using various spectroscopic techniques and DFT methods.     

Application of micro-CT in small animal imaging

Over the past decade, the number of publications using micro-computed tomography (μCT) imaging in preclinical in vivo studies has risen exponentially. Higher spatial and temporal resolution are the key technical advancements that have allowed researchers to capture increasingly detailed anatomical images of small animals and to monitor the progression of disease in small animal models. The purpose of this review is to present the technical aspects of μCT, as well as current research applications. Our objectives are threefold: to familiarize the reader with the basics of μCT techniques; to present the type of experimental designs currently used; and to highlight limitations, future directions, in μCT-scanner research applications, and experimental methods. As a first step we present different μCT setups and components, as well as image contrast generation principles. We then present experimental approaches in order of the evaluated organ system. Finally, we provide a short summary of some of the technical limitations of μCT imaging and discuss potential future developments in μCT-scanner techniques and experimental setups.