A mechanistic study on the tolerance of PAM distal end mismatch by SpCas9.

The therapeutic application of CRISPR-Cas9 is limited due to its off-target activity. To have a better understanding of this off-target effect, we focused on its mismatch-prone PAM distal end. The off-target activity of SpCas9 depends directly on the nature of mismatches, which in turn results in deviation of the active site of SpCas9 due to structural instability in the RNA-DNA duplex strand. In order to test the hypothesis, we designed an array of mismatched target sites at the PAM distal end and performed in vitro and cell line-based experiments, which showed a strong correlation for Cas9 activity. We found that target sites having multiple mismatches in the 18th to 15th position upstream of the PAM showed no to little activity. For further mechanistic validation, Molecular Dynamics simulations were performed, which revealed that certain mismatches showed elevated root mean square deviation values that can be attributed to conformational instability within the RNA-DNA duplex. Therefore, for successful prediction of the off-target effect of SpCas9, along with complementation-derived energy, the RNA-DNA duplex stability should be taken into account.

Unveiling the epigenetic landscape of plants using flow cytometry approach.

Plants are sessile creatures that have to adapt constantly changing environmental circumstances. Plants are subjected to a range of abiotic stressors as a result of unpredictable climate change. Understanding how stress-responsive genes are regulated can help us better understand how plants can adapt to changing environmental conditions. Epigenetic markers that dynamically change in response to stimuli, such as DNA methylation and histone modifications are known to regulate gene expression. Individual cells or particles' physical and/or chemical properties can be measured using the method known as flow cytometry. It may therefore be used to evaluate changes in DNA methylation, histone modifications, and other epigenetic markers, making it a potent tool for researching epigenetics in plants. We explore the use of flow cytometry as a technique for examining epigenetic traits in this thorough discussion. The separation of cell nuclei and their subsequent labeling with fluorescent antibodies, offering information on the epigenetic mechanisms in plants when utilizing flow cytometry. We also go through the use of high-throughput data analysis methods to unravel the complex epigenetic processes occurring inside plant systems.

Persistent chronic calcific pancreatitis with intraductal calculi associated with secondary diabetes mellitus type 3 and diabetic ketoacidosis - A case report.

Diabetes mellitus type 3 refers to diabetes secondary to an existing disease or condition of the exocrine pancreas and is an uncommon cause of diabetes occurring due to pancreatogenic pathology. It accounts for 15-20% of diabetic patients in Indian and Southeast Asian continents. This is case report of a rare case of type 3 diabetes mellitus (T3DM) presenting with diabetic ketoacidosis (DKA). The patient was admitted for DKA along with complaint of hyperglycemia, blood glucose of 405 mg/dl with HbA1c level of 13.7%. Computed tomography evidence revealed chronic calcific pancreatitis with intraductal calculi and dilated pancreatic duct.

Non-potable water reuse and the public health risks from protozoa and helminths: a case study from a city with a semi-arid climate.

The study estimated the risk due to Cryptosporidium, Giardia, and Ascaris, associated with non-potable water reuse in the city of Jaipur, India. The study first determined the exposure dose of Cryptosporidium, Giardia, and Ascaris based on various wastewater treatment technologies for various scenarios of reuse for six wastewater treatment plants (WWTPs) in the city. The exposure scenarios considered were (1) garden irrigation; (2) working and lounging in the garden; and (3) consumption of crops irrigated with recycled water. The estimated annual risk of infection varied between 8.57 × 10-7 and 1.0 for protozoa and helminths, respectively. The order of treatment processes, in decreasing order of annual risk of infection, was found to be: moving-bed bioreactor (MBBR) technology > activated sludge process (ASP) technology > sequencing batch reactor (SBR) technology. The estimated annual risk was found to be in this order: Ascaris > Giardia > Cryptosporidium. The study also estimated the maximum allowable concentration (Cmax) of pathogen in the effluent for a benchmark value of annual infection of risk equal to 1:10,000, the acceptable level of risk used for drinking water. The estimated Cmax values were found to be 6.54 × 10-5, 1.37 × 10-5, and 2.89 × 10-6 (oo) cysts/mL for Cryptosporidium, Giardia, and Ascaris, respectively.

Genome-Wide Meta-Analysis of QTLs Associated with Root Traits and Implications for Maize Breeding.

Root system architecture (RSA), also known as root morphology, is critical in plant acquisition of soil resources, plant growth, and yield formation. Many QTLs associated with RSA or root traits in maize have been identified using several bi-parental populations, particularly in response to various environmental factors. In the present study, a meta-analysis of QTLs associated with root traits was performed in maize using 917 QTLs retrieved from 43 mapping studies published from 1998 to 2020. A total of 631 QTLs were projected onto a consensus map involving 19,714 markers, which led to the prediction of 68 meta-QTLs (MQTLs). Among these 68 MQTLs, 36 MQTLs were validated with the marker-trait associations available from previous genome-wide association studies for root traits. The use of comparative genomics approaches revealed several gene models conserved among the maize, sorghum, and rice genomes. Among the conserved genomic regions, the ortho-MQTL analysis uncovered 20 maize MQTLs syntenic to 27 rice MQTLs for root traits. Functional analysis of some high-confidence MQTL regions revealed 442 gene models, which were then subjected to in silico expression analysis, yielding 235 gene models with significant expression in various tissues. Furthermore, 16 known genes viz., DXS2, PHT, RTP1, TUA4, YUC3, YUC6, RTCS1, NSA1, EIN2, NHX1, CPPS4, BIGE1, RCP1, SKUS13, YUC5, and AW330564 associated with various root traits were present within or near the MQTL regions. These results could aid in QTL cloning and pyramiding in developing new maize varieties with specific root architecture for proper plant growth and development under optimum and abiotic stress conditions.

Glucogallin Attenuates the LPS-Induced Signaling in Macrophages and Protects Mice against Sepsis.

The anti-oxidant and anti-inflammatory effect of beta-glucogallin (BGG), a plant-derived natural product, was evaluated in both in vitro and in vivo studies. For the in vitro study, the ability of BGG pre-treatment to quench LPS-induced effects compared to LPS alone in macrophages was investigated. It was found that BGG pre-treatment showed a significant decrease in ROS, NO, superoxide, and pro-inflammatory cytokines (TNF-alpha, IL-4, IL-17, IL-1ß, and IL-6) and increased reduced glutathione coupled with the restoration of mitochondrial membrane potential. Gene profiling and further validation by qPCR showed that BGG pre-treatment downregulated the LPS-induced expression of c-Fos, Fas, MMP-9, iNOS, COX-2, MyD88, TRIF, TRAF6, TRAM, c-JUN, and NF-κB. We observed that BGG pre-treatment reduced nuclear translocation of LPS-activated NF-κB and thus reduced the subsequent expressions of NLRP3 and IL-1ß, indicating the ability of BGG to inhibit inflammasome formation. Molecular docking studies showed that BGG could bind at the active site of TLR4. Finally, in the LPS-driven sepsis mouse model, we showed that pre-treatment with BGG sustained toxic shock, as evident from their 100% survival. Our study clearly showed the therapeutic potential of BGG in toxic shock syndrome.

In Vitro Anticancer Activity of Methanolic Extract of Justicia adhatoda Leaves with Special Emphasis on Human Breast Cancer Cell Line.

Natural products are being targeted as alternative anticancer agents due to their non-toxic and safe nature. The present study was conducted to explore the in vitro anticancer potential of Justicia adhatoda (J. adhatoda) leaf extract. The methanolic leaf extract was prepared, and the phytochemicals and antioxidant potential were determined by LCMS analysis and DPPH radical scavenging assay, respectively. A docking study performed with five major alkaloidal phytoconstituents showed that they had a good binding affinity towards the active site of NF-κB. Cell viability assay was carried out in five different cell lines, and the extract exhibited the highest cytotoxicity in MCF-7, a breast cancer cell line. Extract-treated cells showed a significant increase in nitric oxide and reactive oxygen species production. Cell cycle analysis showed an arrest in cell growth at the Sub-G0 phase. The extract successfully inhibited cell migration and colony formation and altered mitochondrial membrane potential. The activities of superoxide dismutase and glutathione were also found to decrease in a dose-dependent manner. The percentage of apoptotic cells was found to increase in a dose-dependent manner in MCF-7 cells. The expressions of caspase-3, Bax, and cleaved-PARP were increased in extract-treated cells. An increase in the expression of NF-κB was found in the cytoplasm in extract-treated cells. J. adhatoda leaf extract showed a potential anticancer effect in MCF-7 cells.

Glucogallin Attenuates RAW 264.7 Cells from Arsenic Trioxide Induced Toxicity via the NF-Ò¡B/NLRP3 Pathway.

Chronic arsenic (As) poisoning is mostly due to subsoil water contaminated with As and its salts. Exposure to As has been found to cause an elevation in reactive oxygen species (ROS), leading to the damage of DNA and proteins, and it also causes immunotoxicity. Treatment regimens are primarily based on chelation therapy and amino acid and vitamin supplementations. Recent studies have established that natural products display effective and progressive relief from arsenicosis without any side effects. ß-glucogallin (BGG), a gallo-tannin natural product, is reported to possess anti-oxidant and anti-inflammatory properties. In the present study, we aim to observe the protective role of BGG against As-induced cytotoxicity, apoptosis, mitochondrial dysfunction, and the underlying mechanisms in RAW 264.7 macrophage cells. We found that BGG alleviates As-induced ROS, apoptosis, and mitochondrial dysfunction in RAW 264.7 macrophage cells. Thus, BGG can be used therapeutically to prevent As-induced toxicity.

Carbamazepine versus levetiracetam in epilepsy due to neurocysticercosis.

BACKGROUND: The choice of antiepileptic drug (AED) in newly diagnosed neurocysticercosis (NCC) patients with epilepsy continues to be arbitrary. We compared efficacy and side effect profile of levetiracetam (LEV) and carbamazepine (CBZ) for the treatment of seizures in newly diagnosed patients with NCC. PATIENTS AND METHODS: This was an open-labeled randomized comparative monotherapy study including newly diagnosed drug naïve patients of NCC (n = 99) presenting with seizures who were randomized in 1:1 ratio using computed generated numbers. All patients were followed up for at least six months after start of treatment. The primary outcome measure was seizure control over six months following start of AEDs. RESULTS: Fifteen (15.2%) patients [CBZ- 4(8.2%); LEV- 11(22%)] developed recurrence of seizures. A trend (p = 0.09) was found toward better control of seizures in CBZ compared to LEV. Two (4%) patients in LEV group and 17 (34.6%) patients in CBZ group developed drug-related minor side effects (p < 0.0001). Three patients in CBZ group needed discontinuation of therapy due to skin rash. Eleven patients who relapsed while on LEV did not have any recurrence of seizures after switching over to CBZ. Out of 3 patients who relapsed while receiving CBZ and were changed to LEV, two developed seizures during follow-up. CONCLUSION: CBZ and LEV could be used as alternatives in newly diagnosed patients of NCC at the behest of minor side effects in the CBZ group.

Protease Inhibitory Effect of Natural Polyphenolic Compounds on SARS-CoV-2: An In Silico Study.

The current pandemic, caused by SARS-CoV-2 virus, is a severe challenge for human health and the world economy. There is an urgent need for development of drugs that can manage this pandemic, as it has already infected 19 million people and led to the death of around 711,277 people worldwide. At this time, in-silico studies are providing lots of preliminary data about potential drugs, which can be a great help in further in-vitro and in-vivo studies. Here, we have selected three polyphenolic compounds, mangiferin, glucogallin, and phlorizin. These compounds are isolated from different natural sources but share structural similarities and have been reported for their antiviral activity. The objective of this study is to analyze and predict the anti-protease activity of these compounds on SARS-CoV-2main protease (Mpro) and TMPRSS2 protein. Both the viral protein and the host protein play an important role in the viral life cycle, such as post-translational modification and viral spike protein priming. This study has been performed by molecular docking of the compounds using PyRx with AutoDock Vina on the two aforementioned targets chosen for this study, i.e., SARS-CoV-2 Mpro and TMPRSS2. The compounds showed good binding affinity and are further analyzed by (Molecular dynamic) MD and Molecular Mechanics Poisson-Boltzmann Surface Area MM-PBSA study. The MD-simulation study has predicted that these natural compounds will have a great impact on the stabilization of the binding cavity of the Mpro of SARS-CoV-2. The predicted pharmacokinetic parameters also show that these compounds are expected to have good solubility and absorption properties. Further predictions for these compounds also showed no involvement in drug-drug interaction and no toxicity.

Full Parametric Study of the Influence of Ionomer Content, Catalyst Loading and Catalyst Type on Oxygen and Ion Transport in PEM Fuel Cell Catalyst Layers.

To advance the technology of polymer electrolyte membrane fuel cells, material development is at the forefront of research. This is especially true for membrane electrode assembly, where the structuring of its various layers has proven to be directly linked to performance increase. In this study, we investigate the influence of the various ingredients in the cathode catalyst layer, such as ionomer content, catalyst loading and catalyst type, on the oxygen and ion transport using a full parametric analysis. Using two types of catalysts, 40 wt.% Pt/C and 60 wt.% Pt/C with high surface area carbon, the ionomer/carbon content was varied between 0.29-1.67, while varying the Pt loading in the range of 0.05-0.8 mg cm-2. The optimum ionomer content was found to be dependent on the operating point and condition, as well as catalyst loading and type. The data set provided in this work gives a starting point to further understanding of structured catalyst layers.

The Eye Opener: Finding and Targeting the Midbrain Lesion.

Lesions of the central nucleus of the third cranial nerve in midbrain leads to bilateral ptosis. We present a case of a 14 year old boy, who was found to have B/L non-correctable ptosis and medial rectus palsy. There was a ring enhancing lesion in the midbrain in contrast MRI which resolved with treatment. The case exemplifies that bilateral ptosis results from the central nucleus involvement of third nerve which has bilateral innervation.

Cerebellar ataxia due to Leptospirosis- a case report.

BACKGROUND: Leptospirosis involves nervous system in around 10-15% of the cases, the commonest presentation being aseptic meningitis. Most of the clinical features of neuroleptospirosis are due to capillary endothelial damage and vasculitis. Ataxia is an extremely uncommon manifestation of Leptospirosis occuring in <5% of cases. CASE PRESENTATION: A 28 year old female from North India presented with a short febrile illness followed by an acute onset cerebellar ataxia, anemia, thrombocytopenia and transaminitis. Leptospira serology showed high titres of IgM (ELISA) and MAT (microscopic agglutination test titre >1:800) . She was treated with intravenous ceftriaxone for 14 days following which she showed marked recovery. CONCLUSION: The clinical features of neuroleptospirosis are varied, most of them resulting from endothelial damage and vasculitis. Immune mediated phenomenon with no structural damage is another possible mechanism leading to cerebellar ataxia. Cerebellar ataxia due to common tropical infections should be ruled out in the appropriate setting, as early institution of treatment can abate neurological morbidity. The case report highlights the importance of identifying a reversible cause of cerebellar ataixa due to a tropical infection, possibly due to a immune mediated phenomenon, and would be of interest to both internists and neurologists.

Metabolism-Induced CaCO3 Biomineralization During Reactive Transport in a Micromodel: Implications for Porosity Alteration.

The ability of Pseudomonas stutzeri strain DCP-Ps1 to drive CaCO3 biomineralization has been investigated in a microfluidic flowcell (i.e., micromodel) that simulates subsurface porous media. Results indicate that CaCO3 precipitation occurs during NO3(-) reduction with a maximum saturation index (SIcalcite) of ∼1.56, but not when NO3(-) was removed, inactive biomass remained, and pH and alkalinity were adjusted to SIcalcite ∼ 1.56. CaCO3 precipitation was promoted by metabolically active cultures of strain DCP-Ps1, which at similar values of SIcalcite, have a more negative surface charge than inactive strain DCP-Ps1. A two-stage NO3(-) reduction (NO3(-) → NO2(-) → N2) pore-scale reactive transport model was used to evaluate denitrification kinetics, which was observed in the micromodel as upper (NO3(-) reduction) and lower (NO2(-) reduction) horizontal zones of biomass growth with CaCO3 precipitation exclusively in the lower zone. Model results are consistent with two biomass growth regions and indicate that precipitation occurred in the lower zone because the largest increase in pH and alkalinity is associated with NO2(-) reduction. CaCO3 precipitates typically occupied the entire vertical depth of pores and impacted porosity, permeability, and flow. This study provides a framework for incorporating microbial activity in biogeochemistry models, which often base biomineralization only on SI (caused by biotic or abiotic reactions) and, thereby, underpredict the extent of this complex process. These results have wide-ranging implications for understanding reactive transport in relevance to groundwater remediation, CO2 sequestration, and enhanced oil recovery.

Immobilization of selenite via two parallel pathways during in situ bioremediation.

It is widely understood that selenite can be biologically reduced to elemental selenium. Limited studies have shown that selenite can also be immobilized through abiotic precipitation with sulfide, a product of biological sulfate reduction. We demonstrate that both pathways significantly contribute to selenite immobilization in a microfluidic flow cell having a transverse mixing zone between propionate and selenite that mimics the reaction zone along the margins of a selenite plume undergoing bioremediation in the presence of background sulfate. The experiment showed that red particles of amorphous elemental selenium precipitate on the selenite-rich side of the mixing zone, while long crystals of selenium sulfides precipitate on the propionate-rich side of the mixing zone. We developed a continuum-scale reactive transport model that includes both pathways. The simulated results are consistent with the experimental results, and indicate that spatial segregation of the two selenium precipitates is due to the segregation of the more thermodynamic favorable selenite reduction and the less thermodynamically favorable sulfate reduction. The improved understanding of selenite immobilization and the improved model can help to better design in situ bioremediation processes for groundwater contaminated by selenite or other contaminants (e.g., uranium(IV)) that can be immobilized via similar pathways.

Flow cytometry: Aspects and application in plant and biological science.

Flow cytometry is a potent method that enables the quick and concurrent investigation of several characteristics of single cells in solution. Photodiodes or photomultiplier tubes are employed to detect the dispersed and fluorescent light signals that are produced by the laser beam as it passes through the cells. Photodetectors transform the light signals produced by the laser into electrical impulses. A computer then analyses these electrical impulses to identify and measure the various cell populations depending on their fluorescence or light scattering characteristics. Based on their fluorescence or light scattering properties, cell populations can be examined and/or isolated. This review covers the basic principle, components, working and specific biological applications of flow cytometry, including studies on plant, cell and molecular biology and methods employed for data processing and interpretation as well as the potential future relevance of this methodology in light of retrospective analysis and recent advancements in flow cytometry.

3,5-Disubstituted-thiazolidine-2,4-dione hybrids as antidiabetic agents: Design, synthesis, in-vitro and In vivo evaluation.

Diabetes is one of the fastest-growing metabolic disorders, nearly doubling the number of patients each year. There are different treatment approaches available for the management of diabetes, which lacks due to their side effects. The inhibition of enzymes involved in the metabolism of complex polysaccharides to monosaccharides has proven beneficial in patients with type 2 diabetes mellitus. Two enzymes, α-amylase and α-glucosidase, have emerged as potential drug targets and are widely explored for drug development against type 2 diabetes mellitus. In this context, thiazolidine-2,4-diones (TZDs) have emerged as potential drug candidates for developing newer molecules against α-amylase and α-glucosidase. Nineteen TZD-hybrids were synthesized and evaluated in vitro α-amylase and α-glucosidase inhibitory activity. The compounds 7i, 7k, and 7p have emerged as the best dual inhibitors with IC50 of 10.33 ± 0.11-20.94 ± 0.76 µM and 10.19 ± 0.25-24.07 ± 1.56 µM against α-glucosidase and α-amylase, respectively. The derivatives had good anti-oxidant activity, displaying IC50 = 14.95 ± 0.65-23.27 ± 0.99 µM. The compounds 7k and 7p showed the best inhibition of reactive oxygen species in the PNAC-1 cells. The molecules exhibit good binding within the active site of α-amylase (PDB id: 1B2Y) and α-glucosidase (PDB id: 3W37), displaying binding energies of -7.5 to -10.7 kcal/mol and -7.4 to -10.3 kcal/mol, respectively. Further, the compounds were nontoxic (LD50 = 500-1311 mg/kg) and possessed good GI absorption. The compounds 7i, 7k, and 7p were evaluated in vivo antidiabetic activity in an STZ-induced diabetic model in Wistar rats. The compound 7p emerged as the best compound in the in vivo studies; however, the activity was lesser than that of the standard drug pioglitazone.

A Cross-Sectional Study on Neuropathic Pain Associated With Quality of Sleep in Spondylosis Patients.

INTRODUCTION: Neuropathic pain (NP) is common in spondylosis patients. Cervical and lumbar spondylosis are more common in the elderly population. Spondylosis patients also suffer from poor quality of sleep (QOS). This study aims to find a correlation between NP and QOS in spondylosis patients. METHODS: We conducted a cross-sectional study and analyzed data using the chi-square test to correlate the NP with QOS. The Pittsburgh Sleep Quality Index (PSQI) and the Leeds Assessment of Neuropathic Symptoms and Signs, Self-complete (S-LANSS) questionnaires were used to assess QOS and evaluate neuropathic pain, respectively. Spondylosis was diagnosed based on the history, clinical examination, and radiological findings. RESULTS: A total of 72 spondylosis patients, with a mean age of 47.35 years, were included in this study. Out of 72 subjects, 52 (72.2%) patients had neuropathic pain (NP group), and 20 (27.8%) patients had non-neuropathic pain (non-NP group). In the NP group, 41 patients (78.8%) had poor QOS, while 11 (21.2%) had good QOS. In the non-NP group, eight (40%) had poor QOS, and 12 (60%) had good QOS. CONCLUSION: This study concludes that neuropathic pain is associated with poor quality of sleep in spondylosis patients.

Designing of xanthine-based DPP-4 inhibitors: a structure-guided alignment dependent Multifacet 3D-QSAR modeling, and molecular dynamics simulation study.

The DPP-4 enzyme degrades incretin hormones GLP-1 and GIP. DPP-4 inhibitors are found effective in the prevention of the degradation of incretins. Xanthine scaffold-bearing molecules are reported as potential DPP-4 inhibitors for treating type 2 diabetes mellitus, e.g. the marketed drug linagliptin. In this work, structure-guided alignment-dependent atom- and Gaussian field-based 3D-QSAR have been performed on a dataset of 75 molecules. The robustness and predictive ability of the developed multifacet 3D-QSAR models were validated on different statistical parameters and found to be statistically fit. The favorable and unfavorable pharmacophoric features were mapped for each multifacet 3D-QSAR model based on three alignment sets (1-3). A five-point common pharmacophore hypothesis was generated separately for each set of alignments. The molecular dynamics simulations (up to 100 ns) were performed for the potent molecule from each alignment set (Compounds 12, 40 and 57) compared to reference standard linagliptin to study the binding energy and stability of target-ligand complexes. The MM-PBSA calculations revealed that the binding free energy and stability of compounds 12 (-40.324 ± 17.876 kJ/mol), 40 (-80.543 ± 21.782 kJ/mol) and 57 (-50.202 ± 16.055 kJ/mol) were better than the reference drug linagliptin (-20.390 ± 63.200 kJ/mol). The generated contour maps from structure-guided alignment-dependent multifacet 3D-QSAR models offer information about the structure-activity relationship (SAR) and ligand-target binding energy and stability data from MD simulation may be utilized to design and develop target selective xanthine-based novel DPP-4 inhibitors.Communicated by Ramaswamy H. Sarma.

Multistage in silico approach to identify novel quinoline derivatives as potential c-kit kinase inhibitors.

The type II-C-KIT signaling network has been extensively studied for its potential as a target for cancer treatment, leading to the investigation of quinoline derivatives as compounds with inhibitory effects on c-Kit kinase. In this study, a multistage approach was employed, including the creation of pharmacophore models, 3D QSAR analysis, virtual screening, docking investigations, and molecular dynamics stimulation. The pharmacophore evaluation included a data set of 29 ligands, which resulted in the generation of the ADDHR_1pharmacophore model as the most promising, with a survival score of 5.6812. The main objective was to utilize the atom-based 3D-QSAR approach for generating robust 3D-QSAR models aimed at identifying new TypeII-C-kit kinase inhibitors. The evaluations of these models have convincingly demonstrated their high predictive power (Q2 = 0.6547, R2 = 0.9947). Using atom-based 3D-QSAR data, a total of 7564 novel compounds were generated from R-group enumeration. Molecular docking and MM-GBSA study revealed that compound A1 exhibited the highest binding score of -9.30 kcal/mol and a Δ GBind value of -90.56 kcal/mol. The ZINC compounds were then screened using the pharmacophore model, followed by virtual screening, which identified ZINC65798256, ZINC09317958, ZINC73187176, and ZINC76176670 as potential candidates with promising docking scores. Among these, ZINC65798256 demonstrated the best binding interactions with amino acid residues, ASP810, LYS623, CYS673, and THR670 (PDB ID: 1T46). To further analyze the structural features and molecular interactions, molecular dynamics simulation was conducted for a time scale of 100 ns.Communicated by Ramaswamy H. Sarma.