Multispectroscopic insight,morphological analysis and molecular docking studies of CuII-based chemotherapeutic drug entity with human serum albumin (HSA) and bovine serum albumin (BSA)

The interaction studies of Cu^II nalidixic acid–DACH chemotherapeutic drug entity, [C₃₆H₅₀N₈O₆Cu] with serum albumin proteins, viz., human serum albumin (HSA) and bovine serum albumin (BSA) employing UV–vis, fluorescence, CD, FTIR and molecular docking techniques have been carried out. Complex [C₃₆H₅₀N₈O₆Cu] demonstrated strong binding affinity towards serum albumin proteins via hydrophobic contacts with binding constants, K?=?3.18?×?10⁵ and 7.44?×?10⁴ M^–1 for HSA and BSA, respectively implicating a higher binding affinity for HSA. The thermodynamic parameters ΔG, ΔH and ΔS at different temperatures were also calculated and the interaction of complex [C₃₆H₅₀N₈O₆Cu] with HSA and BSA was found to be enthalpy and entropy favoured, nevertheless, complex [C₃₆H₅₀N₈O₆Cu] demonstrated higher binding affinity towards HSA than BSA evidenced from its higher binding constant values. Time resolved fluorescence spectroscopy (TRFS) was carried out to validate the static quenching mechanism of HSA/BSA fluorescence. The collaborative results of spectroscopic studies indicated that the microenvironment and the conformation of HSA and BSA (α–helix) were significantly perturbed upon interaction with complex [C₃₆H₅₀N₈O₆Cu]. Hirshfeld surfaces analysis and fingerprint plots revealed various intermolecular interactions viz., N–H····O, O–H····O and C–H····O linkages in a 2–dimensional framework that provide crucial information about the supramolecular architectures in the complex. Molecular docking studies were carried out to ascertain the preferential binding mode and affinity of complex [C₃₆H₅₀N₈O₆Cu] at the target site of HSA and BSA. Furthermore, only for Transmission electroscopy microscopy micrographs of HSA and BSA in presence of complex [C₃₆H₅₀N₈O₆Cu] revealed major protein morphological transitions and aggregation which validates efficient delivery of complex by serum proteins to the target site.

Communicated by Ramaswamy H. Sarma     

Predictive equations to estimate spinal loads in symmetric lifting tasks

Response surface methodology is used to establish robust and user-friendly predictive equations that relate responses of a complex detailed trunk finite element biomechanical model to its input variables during sagittal symmetric static lifting activities. Four input variables (thorax flexion angle, lumbar/pelvis ratio, load magnitude, and load position) and four model responses (L4–L5 and L5–S1 disc compression and anterior–posterior shear forces) are considered. Full factorial design of experiments accounting for all combinations of input levels is employed. Quadratic predictive equations for the spinal loads at the L4–S1 disc mid-heights are obtained by regression analysis with adequate goodness-of-fit (R²>98%, p<0.05, and low root-mean-squared-error values compared with the range of predicted spine loads). Results indicate that intradiscal pressure values at the L4–L5 disc estimated based on the predictive equations are in close agreement with available in vivo data measured under similar loadings and postures. Combinations of input (posture and loading) variable levels that yield spine loads beyond the tolerance compression limit of 3400 N are identified using contour plots. Ergonomists and bioengineers, faced with the dilemma of using either complex but more accurate models on one hand or less accurate but simple models on the other hand, have thereby easy-to-use predictive equations that quantifies spinal loads and risk of injury under different occupational tasks of interest.     

Cyclin-dependent kinase inhibitors,roscovitine and purvalanol,induce apoptosis and autophagy related to unfolded protein response in HeLa cervical cancer cells

Roscovitine (Rosc) and purvalanol (Pur) are competitive inhibitors of cyclin-dependent kinases (CDKs) by targeting their ATP-binding pockets. Both drugs are shown to be effective to decrease cell viability and dysregulate the ratio of pro- and anti-apoptotic Bcl-2 family members, which finally led to apoptotic cell death in different cancer cell lines in vitro. It was well established that Bcl-2 family members have distinct roles in the regulation of other cellular processes such as endoplasmic reticulum (ER) stress. The induction of ER stress has been shown to play critical role in cell death/survival decision via autophagy or apoptosis. In this study, our aim was to investigate the molecular targets of CDK inhibitors on ER stress mechanism related to distinct cell death types in time-dependent manner in HeLa cervical cancer cells. Our results showed that Rosc and Pur decreased the cell viability, cell growth and colony formation, induced ER stress-mediated autophagy or apoptosis in time-dependent manner. Thus, we conclude that exposure of cells to CDK inhibitors induces unfolded protein response and ER stress leading to autophagy and apoptosis processes in HeLa cervical cancer cells.     

Labelling of lupine nodule metabolites with 14CO2 assimilated from the leaves

On feeding ¹⁴CO₂ to the shoots of lupine (25 mCi per plant) 30 min was the minimal time needed to determine the incorporation of label into bacteroid compounds. The predominant incorporation, exhibited in all root, nodule and bacteroid samples after 30 min exposure, was into sucrose (45–90% of the corresponding fraction radioactivity) of the neutral fraction; into malate (30–40%) of the acid fraction; into aspartic acid and asparagine (60–80% in sum) of the basic fraction. The composition of carbon compounds containing the greatest amount of ¹⁴C in the cytosol of nodules and in bacteroids was similar. Their radioactivity after 30 min exposure was for bacteroids (nCi per g of bacteroid fr. wt): sucrose 5.73, glucose 1.00, malate 0.15, succinate 0.11; for the nodule cytosol (nCi per g of nodule fr. wt): sucrose 200.00, glucose 8.40, malate 9.34, succinate 8.50. Thus it was demonstrated that in lupine, sucrose is the main photoassimilate entering not only into nodules but also into bacteroids. The biosynthesis of aspartic acid and asparagine occurs during nitrogen fixation in bacteroids.     

In vivo effects of curcumin on the paraoxonase,carbonic anhydrase,glucose-6-phosphate dehydrogenase and β-glucosidase enzyme activities in dextran sulphate sodium-induced ulcerative colitis mice

Increases in the risk of infections and malignancy due to immune suppressive therapies of inflammatory bowel diseases (IBDs) have led the researchers to focus on more nontoxic and acceptable natural products like curcumin. Here we investigate whether prophylactic and therapeutic application of the curcumin alters the enzyme activities of paraoxonase (PON), carbonic anhydrase (CA), glucose-6-phosphate dehydrogenase (G6PD) and cytosolic β-glucosidase in dextran sulphate sodium (DSS)-induced ulcerative colitis mice. Prophylactic application of curcumin resulted in higher MPO activity, less body weight loss and longer colon lengths compared to therapeutic group indicating preventive role of curcumin in IBDs. DSS-induced decrease in liver and serum PON activities were completely recovered by prophylactic administration of curcumin. DSS-induced reduction in liver cytosolic β-glucosidase activity was not affected by curcumin neither in the prophylactic group nor in the therapeutic group. Erythrocyte CA activity was significantly increased in curcumin groups, however no remarkable change in G6PD activity was observed.     

Intracellular Gating in an Inward-facing State of Aspartate Transporter GltPh Is Regulated by the Movements of the Helical Hairpin HP2

Sodium-coupled neurotransmitter transporters play a key role in neuronal signaling by clearing excess transmitter from the synapse. Structural data on a trimeric archaeal aspartate transporter, Glt_Ph, have provided valuable insights into structural features of human excitatory amino acid transporters. However, the time-resolved mechanisms of substrate binding and release, as well as that of coupling to sodium co-transport, remain largely unknown for this important family. We present here the results of the most extensive simulations performed to date for Glt_Ph in both outward-facing and inward-facing states by taking advantage of significant advances made in recent years in molecular simulation technology. The generated multiple microsecond trajectories consistently show that the helical hairpin HP2, not HP1, serves as an intracellular gate (in addition to its extracellular gating role). In contrast to previous proposals, HP1 can neither initiate nor accommodate neurotransmitter release without prior opening of HP2 by at least 4.0 Å. Aspartate release invariably follows that of a sodium ion located near the HP2 gate entrance. Asp-394 on TM8 and Arg-276 on HP1 emerge as key residues that promote the reorientation and diffusion of substrate toward the cell interior. These findings underscore the significance of examining structural dynamics, as opposed to static structure(s), to make inferences on the mechanisms of transport and key interactions.     

Coupled Global and Local Changes Direct Substrate Translocation by?Neurotransmitter-Sodium Symporter Ortholog LeuT

Significant advances have been made in recent years in characterizing neurotransmitter:sodium symporter (NSS) family structure and function. Yet, many time-resolved events and intermediates that control the various stages of transport cycle remain to be elucidated. Whether NSSs harbor one or two sites for binding their substrates (neurotransmitters or amino acids), and what the role of the secondary site S2 is, if any, are still unresolved. Using molecular modeling and simulations for LeuT, a bacterial NSS, we present a comprehensive account of substrate-binding and -stabilization events, and subsequently triggered interactions leading to substrate (alanine) release. LeuT instantaneous conformation as it reconfigures from substrate-receiving (outward-facing) to -releasing (inward-facing) state appears to be a determinant of its affinity to bind substrate at site S2. In the outward-facing state, S1 robustly binds alanine and regulates subsequent redistribution of interactions to trigger extracellular gate closure; whereas S2 is only a transient binding site. The substrate-binding affinity at S2 increases in an intermediate close to inward-facing state. LeuT harbors the two substrate-binding sites, and small displacements of second substrate near S2 are observed to induce concerted small translocations in the substrate bound to primary site S1, although complete release requires collective structural rearrangements that fully expose the intracellular vestibule to the cytoplasm.     

Recruitment of rare <Emphasis Type="Italic">3</Emphasis>-grams at functional sites: Is this a mechanism for increasing enzyme specificity?

Background  

A wealth of unannotated and functionally unknown protein sequences has accumulated in recent years with rapid progresses in sequence genomics, giving rise to ever increasing demands for developing methods to efficiently assess functional sites. Sequence and structure conservations have traditionally been the major criteria adopted in various algorithms to identify functional sites. Here, we focus on the distributions of the 20³ different types of 3-grams (or triplets of sequentially contiguous amino acid) in the entire space of sequences accumulated to date in the UniProt database, and focus in particular on the rare 3-grams distinguished by their high entropy-based information content.