Molecular dynamic simulations analysis of ritonavir and lopinavir as SARS-CoV 3CL(pro) inhibitors

Since the emergence of the severe acute respiratory syndrome (SARS) to date, neither an effective antiviral drug nor a vaccine against SARS is available. However, it was found that a mixture of two HIV-1 proteinase inhibitors, lopinavir and ritonavir, exhibited some signs of effectiveness against the SARS virus. To understand the fine details of the molecular interactions between these proteinase inhibitors and the SARS virus via complexation, molecular dynamics simulations were carried out for the SARS-CoV 3CL^pro free enzyme (free SARS) and its complexes with lopinavir (SARS-LPV) and ritonavir (SARS-RTV). The results show that flap closing was clearly observed when the inhibitors bind to the active site of SARS-CoV 3CL^pro. The binding affinities of LPV and RTV to SARS-CoV 3CL^pro do not show any significant difference. In addition, six hydrogen bonds were detected in the SARS-LPV system, while seven hydrogen bonds were found in SARS-RTV complex.     

Maternal footprints of Southeast Asians in North India

We have analyzed 7,137 samples from 125 different caste, tribal and religious groups of India and 99 samples from three populations of Nepal for the length variation in the COII/tRNA(Lys) region of mtDNA. Samples showing length variation were subjected to detailed phylogenetic analysis based on HVS-I and informative coding region sequence variation. The overall frequencies of the 9-bp deletion and insertion variants in South Asia were 1.9 and 0.6%, respectively. We have also defined a novel deep-rooting haplogroup M43 and identified the rare haplogroup H14 in Indian populations carrying the 9-bp deletion by complete mtDNA sequencing. Moreover, we redefined haplogroup M6 and dissected it into two well-defined subclades. The presence of haplogroups F1 and B5a in Uttar Pradesh suggests minor maternal contribution from Southeast Asia to Northern India. The occurrence of haplogroup F1 in the Nepalese sample implies that Nepal might have served as a bridge for the flow of eastern lineages to India. The presence of R6 in the Nepalese, on the other hand, suggests that the gene flow between India and Nepal has been reciprocal.     

Integrated Recovery of Pigments Released from Red Beet Hairy Roots Exposed to Acidic Medium

Hairy root cultures of Beta vulgaris L grown in a bubble column reactor were permeabilised by exposure to B5 medium of pH 2.0. The roots released 39% of their total pigments on a 10 min exposure to B5 medium of pH 2.0 followed by return to standard 135 medium. The pigments released in the extracellular medium were recovered on an adsorption column containing XAD-16 resin. The permeabilised roots regrew and accumulated additional pigments. Comparison of this technique with the previously used techniques like oxygen starvation and temperature shock to permeabilise beet hairy roots suggest that pH mediated release of betalains can be an effective method of releasing betalains from root cultures.     

Screening Fluorescent Voltage Indicators with Spontaneously Spiking HEK Cells

Development of improved fluorescent voltage indicators is a key challenge in neuroscience, but progress has been hampered by the low throughput of patch-clamp characterization. We introduce a line of non-fluorescent HEK cells that stably express Na_V 1.3 and K_IR 2.1 and generate spontaneous electrical action potentials. These cells enable rapid, electrode-free screening of speed and sensitivity of voltage sensitive dyes or fluorescent proteins on a standard fluorescence microscope. We screened a small library of mutants of archaerhodopsin 3 (Arch) in spiking HEK cells and identified two mutants with greater voltage-sensitivity than found in previously published Arch voltage indicators.     

Complete chloroplast genome of the medicinal plant Evolvulus alsinoides: comparative analysis,identification of mutational hotspots and evolutionary dynamics with species of Solanales

Evolvulus alsinoides, belonging to the family Convolvulaceae, is an important medicinal plant widely used as a nootropic in the Indian traditional medicine system. In the genus Evolvulus, no research on the chloroplast genome has been published. Hence, the present study focuses on annotation, characterization, identification of mutational hotspots, and phylogenetic analysis in the complete chloroplast genome (cp) of E. alsinoides. Genome comparison and evolutionary dynamics were performed with the species of Solanales. The cp genome has 114 genes (80 protein-coding genes, 30 transfer RNA, and 4 ribosomal RNA genes) that were unique with total genome size of 157,015 bp. The cp genome possesses 69 RNA editing sites and 44 simple sequence repeats (SSRs). Predicted SSRs were randomly selected and validated experimentally. Six divergent hotspots such as trnQ-UUG, trnF-GAA, psaI, clpP, ndhF, and ycf1 were discovered from the cp genome. These microsatellites and divergent hot spot sequences of the Taxa ‘Evolvulus’ could be employed as molecular markers for species identification and genetic divergence investigations. The LSC area was found to be more conserved than the SSC and IR region in genome comparison. The IR contraction and expansion studies show that nine genes rpl2, rpl23, ycf1, ycf2, ycf1, ndhF, ndhA, matK, and psbK were present in the IR-LSC and IR-SSC boundaries of the cp genome. Fifty-four protein-coding genes in the cp genome were under negative selection pressure, indicating that they were well conserved and were undergoing purifying selection. The phylogenetic analysis reveals that E. alsinoides is closely related to the genus Cressa with some divergence from the genus Ipomoea. This is the first time the chloroplast genome of the genus Evolvulus has been published. The findings of the present study and chloroplast genome data could be a valuable resource for future studies in population genetics, genetic diversity, and evolutionary relationship of the family Convolvulaceae.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12298-021-01051-w.     

The role of molecular markers in predicting response to therapy in patients with colorectal cancer

Advances in systemic therapy for colorectal cancer have dramatically improved prognosis. While disease stage has traditionally been the main determinant of disease course, several molecular characteristics of tumor specimens have recently been shown to have prognostic significance. Although to date no molecular characteristics have emerged as consistent predictors of response to therapy, retrospective studies have investigated the role of a variety of biomarkers, including microsatellite instability, loss of heterozygosity of 18q, type II transforming growth factor beta receptor, thymidylate synthase, epidermal growth factor receptor, and Kirsten-ras (KRAS). This paper reviews the current literature, ongoing prospective studies evaluating the role of these markers, and novel techniques such as gene profiling, which may help to uncover the more complex molecular interactions that will predict response to chemotherapy in patients with colorectal cancer.     

Localization of betav tubulin in the cochlea and cultured cells with a novel monoclonal antibody

Tubulin, the dimeric structural protein of microtubules, is a heterodimer of alpha and beta subunits; both alpha and beta exist as numerous isotypes encoded by different genes. In vertebrates the sequence differences among the beta(I), beta(II), beta(III), beta(IV) and beta(V) isotypes are highly conserved in evolution, implying that the isotypes may have functional significance. Isotype-specific monoclonal antibodies have been useful in determining the cellular and sub-cellular distributions and possible functions of the beta(I), beta(II), beta(III), and beta(IV) isotypes; however, little is known about the beta(V) isotype. We here report the creation and purification of a monoclonal antibody (SHM.12G11) specific for beta(V). The antibody was designed to be specific for the C-terminal sequence EEEINE, which is unique to rodent and chicken beta(V). The antibody was found to bind specifically to the C-terminal peptide EEEINE, and does not cross-react with the carboxy-termini of either alpha-tubulin or the other beta-tubulin isotypes. However, the antibody also binds to the peptide EEEVNE, but not to the peptide EEEIDG, corresponding respectively to the C-terminal peptides of bovine and human beta(V). Immunofluorescence analysis indicates that beta(V) is found in microtubules of both the interphase network and the mitotic spindle. In gerbils, beta(V) also occurs in the cochlea where it is found largely in the specialized cells that are unique in containing bundled microtubules with 15 protofilaments.