Crystal Structure of Allophycocyanin from Marine Cyanobacterium Phormidium sp. A09DM

Isolated phycobilisome (PBS) sub-assemblies have been widely subjected to X-ray crystallography analysis to obtain greater insights into the structure-function relationship of this light harvesting complex. Allophycocyanin (APC) is the phycobiliprotein always found in the PBS core complex. Phycocyanobilin (PCB) chromophores, covalently bound to conserved Cys residues of α- and β- subunits of APC, are responsible for solar energy absorption from phycocyanin and for transfer to photosynthetic apparatus. In the known APC structures, heterodimers of α- and β- subunits (known as αβ monomers) assemble as trimer or hexamer. We here for the first time report the crystal structure of APC isolated from a marine cyanobacterium (Phormidium sp. A09DM). The crystal structure has been refined against all the observed data to the resolution of 2.51 Å to R_work (R_free) of 0.158 (0.229) with good stereochemistry of the atomic model. The Phormidium protein exists as a trimer of αβ monomers in solution and in crystal lattice. The overall tertiary structures of α- and β- subunits, and trimeric quaternary fold of the Phormidium protein resemble the other known APC structures. Also, configuration and conformation of the two covalently bound PCB chromophores in the marine APC are same as those observed in fresh water cyanobacteria and marine red algae. More hydrophobic residues, however, constitute the environment of the chromophore bound to α-subunit of the Phormidium protein, owing mainly to amino acid substitutions in the marine protein.     

Local fractal dimension based ECG arrhythmia classification

We propose a local fractal dimension based nearest neighbor classifier for ECG based classification of arrhythmia. Local fractal dimension (LFD) at each sample point of the ECG waveform is taken as the feature. A nearest neighbor algorithm in the feature space is used to find the class of the test ECG beat. The nearest neighbor is found based on the RR-interval-information-biased Euclidean distance, proposed in the current work. Based on the two algorithms used for estimating the LFD, two classification algorithms are validated in the current work, viz. variance based fractal dimension estimation based nearest neighbor classifier and power spectral density based fractal dimension estimation based nearest neighbor classifier. Their performances are evaluated based on various figures of merit. MIT-BIH (Massachusetts Institute of Technology - Boston’s Beth Israel Hospital) Arrhythmia dataset has been used to validate the algorithms. Along with showing good performance against all the figures of merit, the proposed algorithms also proved to be patient independent in the sense that the performance is good even when the test ECG signal is from a patient whose ECG is not present in the training ECG dataset.     

Ratiometric imaging of calcium during ischemiareperfusion injury in isolated mouse hearts using Fura-2

ABSTRACT: BACKGROUND: We present an easily implementable method for measuring Fura-2 fluorescence from isolated mouse hearts using a commercially available switching light source and CCD camera. After calibration, it provides a good estimate of intracellular [Ca2+] with both high spatial and temporal resolutions, permitting study of changes in dispersion of diastolic [Ca2+], Ca2+ transient dynamics, and conduction velocities in mouse hearts. In a proof-of-principle study, we imaged isolated Langendorff-perfused mouse hearts with reversible regional myocardial infarctions. METHODS: Isolated mouse hearts were perfused in the Landendorff-mode and loaded with Fura-2. Hearts were then paced rapidly and subjected to 15 minutes of regional ischemia by ligation of the left anterior descending coronary artery, following which the ligation was removed to allow reperfusion for 15 minutes. Fura-2 fluorescence was recorded at regular intervals using a high-speed CCD camera. The two wavelengths of excitation light were interleaved at a rate of 1 KHz with a computer controlled switching light source to illuminate the heart. RESULTS: Fura-2 produced consistent Ca2+ transients from different hearts. Ligating the coronary artery rapidly generated a well defined region with a dramatic rise in diastolic Ca2+ without a significant change in transient amplitude; Ca2+ handling normalized during reperfusion. Conduction velocity was reduced by around 50% during ischemia, and did not recover significantly when monitored for 15 minutes following reperfusion. CONCLUSIONS: Our method of imaging Fura-2 from isolated whole hearts is capable of detecting pathological changes in intracellular Ca2+ levels in cardiac tissue. The persistent change in the conduction velocities indicates that changes to tissue connectivity rather than altered intracellular Ca2+ handling may be underlying the electrical instabilities commonly seen in patients following a myocardial infarction.     

Magnetic and contrast properties of labeled platelets for magnetomotive optical coherence tomography

This article introduces a new functional imaging paradigm that uses optical coherence tomography (OCT) to detect rehydrated, lyophilized platelets (RL platelets) that are in the preclinical trial stage and contain superparamagnetic iron oxides (SPIOs) approved by the U.S. Food and Drug Administration. Platelets are highly functional blood cells that detect and adhere to sites of vascular endothelial damage by forming primary hemostatic plugs. By applying magnetic gradient forces, induced nanoscale displacements (magnetomotion) of the SPIO-RL platelets are detected as optical phase shifts in OCT. In this article, we characterize the iron content and magnetic properties of SPIO-RL platelets, construct a model to predict their magnetomotion in a tissue medium, and demonstrate OCT imaging in tissue phantoms and ex vivo pig arteries. Tissue phantoms containing SPIO-RL platelets exhibited >3 dB contrast/noise ratio at ≥1.5 × 10⁹ platelets/cm³. OCT imaging was performed on ex vivo porcine arteries after infusion of SPIO-RL platelets, and specific contrast was obtained on an artery that was surface-damaged (P < 10⁻⁶). This may enable new technologies for in vivo monitoring of the adherence of SPIO-RL platelets to sites of bleeding and vascular damage, which is broadly applicable for assessing trauma and cardiovascular diseases.     

Characterization of Parasarcophaga heterochromatin

The heterochromatin of neuroblast metaphase chromosomes in six sympatric species of Parasarcophaga was characterized by C-, N-, quinacrine (Q)-, Hoechst 33258 (H)-, Hoechst/actinomycin D (H/AMD)- and acridine orange (AO)-banding. The autosomes and the sex chromosomes in all the six species are characterized by the presence of C- and N-positive heterochromatin. The N-bands lie within the limits of the C⁺ heterochromatin. AT-specific fluorochromes further differentiate heterochromatin into Q⁺H⁺,Q^–H⁺ and Q^–H^– types and the repetitive nature of these regions is indicated by their fast reassociating nature with AO-staining. Thus, on the basis of these results, the heterochromatin of mitotic chromosomes in Parasarcophaga can be classified into C⁺N⁺Q⁺H⁺, C⁺N⁺Q^–H^–, c⁺n^–q^–h^– and C^–N^–Q^–H⁺ types.     

Direct in vitro regeneration of medicinally important Indian and exotic red-colored ginger (Zingiber officinale Rosc.) and genetic fidelity assessment using ISSR and SSR markers

In Vitro Cellular & Developmental Biology - Plant - An efficient direct in vitro plant regeneration protocol has been established for medicinally important red-colored ginger (Zingiber...     

Plant-associated endophytic fungi as potential bio-factories for extracellular enzymes: Progress,Challenges and Strain improvement with precision approaches

There is an intricate network of relations between endophytic fungi and their hosts that affects the production of various bioactive compounds. Plant-associated endophytic fungi contain industrially important enzymes and have the potential to fulfil their rapid demand in the international market to boost business in technology. Being safe and metabolically active, they have replaced the usage of toxic and harmful chemicals and hold a credible application in biotransformation, bioremediation and industrial processes. Despite these, there are limited reports on fungal endophytes that can directly cater to the demand and supply of industrially stable enzymes. The underlying reasons include low endogenous production and secretion of enzymes from fungal endophytes which have raised concern for widely accepted applications. Hence, it is imperative to augment the biosynthetic and secretory potential of fungal endophytes. Modern state-of-the-art biotechnological technologies aiming at strain improvement using cell factory engineering as well as precise gene editing like Clustered Regularly Interspaced Palindromic Repeats (CRISPR) and its Associated proteins (Cas) systems which can provide a boost in fungal endophyte enzyme production. Additionally, it is vital to characterize optimum conditions to grow one strain with multiple enzymes (OSME). The present review encompasses various plants-derived endophytic fungal enzymes and their applications in various sectors. Furthermore, we postulate the feasibility of new precision approaches with an aim for strain improvement and enhanced enzyme production.     

Prevalence and Characterization of <Emphasis Type="Italic">Cronobacter</Emphasis> spp. from Various Foods,Medicinal Plants,and Environmental Samples

Dairy or non-dairy based products were explored to determine the prevalence, molecular characterization, and antibiotic susceptibility of Cronobacter spp. The isolation was done as per ISO 22964:2006 on chromogenic media followed by further confirmation by biochemical- and 16S rRNA-based identification. From 219 samples, the chromogenic agar assay and biochemical tests yielded presumptive 45 isolates. Among them, only 36 isolates showed 282 bp band amplified from ITS-G gene confirming as Cronobacter sakazakii. The Cronobacter spp. prevalence was highest in herbs and spices (34 %) while environmental samples had contamination rates of 23 % indicating plants as a possible reservoir of this pathogen. All the isolates were resistant to β-lactam derivatives (68 %), macrolides (88.6 %), and aminoglycosides (79.9 %) but susceptible to phenicoles (31.6 %) and tetracyclines (15 %) derivatives. The results emphasize the screening of plant materials before their incorporation in food matrices.     

Cyclosporin A Disrupts Notch Signaling and Vascular Lumen Maintenance

Cyclosporin A (CSA) suppresses immune function by blocking the cyclophilin A and calcineurin/NFAT signaling pathways. In addition to immunosuppression, CSA has also been shown to have a wide range of effects in the cardiovascular system including disruption of heart valve development, smooth muscle cell proliferation, and angiogenesis inhibition. Circumstantial evidence has suggested that CSA might control Notch signaling which is also a potent regulator of cardiovascular function. Therefore, the goal of this project was to determine if CSA controls Notch and to dissect the molecular mechanism(s) by which CSA impacts cardiovascular homeostasis. We found that CSA blocked JAG1, but not Dll4 mediated Notch1 NICD cleavage in transfected 293T cells and decreased Notch signaling in zebrafish embryos. CSA suppression of Notch was linked to cyclophilin A but not calcineurin/NFAT inhibition since N-MeVal-4-CsA but not FK506 decreased Notch1 NICD cleavage. To examine the effect of CSA on vascular development and function, double transgenic Fli1-GFP/Gata1-RFP zebrafish embryos were treated with CSA and monitored for vasculogenesis, angiogenesis, and overall cardiovascular function. Vascular patterning was not obviously impacted by CSA treatment and contrary to the anti-angiogenic activity ascribed to CSA, angiogenic sprouting of ISV vessels was normal in CSA treated embryos. Most strikingly, CSA treated embryos exhibited a progressive decline in blood flow that was associated with eventual collapse of vascular luminal structures. Vascular collapse in zebrafish embryos was partially rescued by global Notch inhibition with DAPT suggesting that disruption of normal Notch signaling by CSA may be linked to vascular collapse. However, multiple signaling pathways likely cause the vascular collapse phenotype since both cyclophilin A and calcineurin/NFAT were required for normal vascular function. Collectively, these results show that CSA is a novel inhibitor of Notch signaling and vascular function in zebrafish embryos.