'Computational studies on coumestrol-ArlR interaction to target ArlRS signaling cascade involved in MRSA virulence'.

Two component signaling system ArlRS (Autolysis-related locus) regulates adhesion, biofilm formation and virulence in methicillin resistant Staphylococcus aureus. It consists of a histidine kinase ArlS and response regulator ArlR. ArlR is composed of a N-terminal receiver domain and DNA-binding effector domain at C-terminal. ArlR receiver domain dimerizes upon signal recognition and activates DNA binding by effector domain and subsequent virulence expression. In silico simulation and structural data suggest that coumestrol, a phytochemical found in Pueraria montana, forges a strong intermolecular interaction with residues involved in dimer formation and destabilizes ArlR dimerization, an essential conformational switch required for downstream effector domain to bind to virulent loci. Structural and energy profiles of simulated ArlR-coumestrol complexes suggest lower affinity between ArlR monomers due to structural rigidity at the dimer interface hindering the conformational rearrangements relevant for dimer formation. These analyses could be an attractive strategy to develop therapeutics and potent leads molecules response regulators of two component systems in which are involved in MRSA virulence as well as other drug-resistant pathogens.Communicated by Ramaswamy H. Sarma.

Structural and molecular dynamics of ammonia transport in Staphylococcus aureus NH3-dependent NAD synthetase.

Ammonia dependent NAD+ synthetase from multi drug resistance Staphylococcus aureus catalyzes ATP dependent formation of NAD+ from deamido-NAD+ and ammonia at the synthetase active site. Binding of ATP accompanies a large movement of flexible loop region (205-225) acting as a lid to the catalytic core. A 17 Å long ammonia tunnel with an entry and exit radius of 3.5 Å and 3.2 Å respectively allows transfer of ammonia from surface to the active site of the enzyme in each monomer to attack the C7N=O7N linkage of transient intermediate NAD-adenylate thus releasing NAD+. In this study, we report structural details of ammonia transport tunnel in Staphylococcus aureus NH3-dependent NAD synthetase and compared their architecture and dynamics with other bacterial and eukaryotic enzymes. Tunnel shows conformational variations in apo and substrate complexes and is less intricate compared to glutamine dependent counterparts. We have also performed steered molecular dynamic simulations of ammonia transport across the tunnel in enzyme-intermediate complex which reveals critical bottleneck residues and structural determinants during ammonium migration. Ordered water molecules and conserved charged residues form a network of hydrogen bonds and electrostatic interaction which facilitate the ammonium movement towards the active center. Analysis of the sMD simulated structural snapshots delineates the conformational reshaping of ammonia tunnel at the different step of the enzymatic reaction. Tunnel architecture and environment could offer the new target site to design novel small molecule inhibitors for the development of more efficient therapeutics against multi drug resistant S. aureus strains.

Bruton's Tyrosine Kinase Targeting in Multiple Myeloma.

Multiple myeloma (MM), a clonal plasma cell disorder, disrupts the bones' hematopoiesis and microenvironment homeostasis and ability to mediate an immune response against malignant clones. Despite prominent survival improvement with newer treatment modalities since the 2000s, MM is still considered a non-curable disease. Patients experience disease recurrence episodes with clonal evolution, and with each relapse disease comes back with a more aggressive phenotype. Bruton's Tyrosine Kinase (BTK) has been a major target for B cell clonal disorders and its role in clonal plasma cell disorders is under active investigation. BTK is a cytosolic kinase which plays a major role in the immune system and its related malignancies. The BTK pathway has been shown to provide survival for malignant clone and multiple myeloma stem cells (MMSCs). BTK also regulates the malignant clones' interaction with the bone marrow microenvironment. Hence, BTK inhibition is a promising therapeutic strategy for MM patients. In this review, the role of BTK and its signal transduction pathways are outlined in the context of MM.

Crystallographic and molecular dynamics simulation analysis of NAD synthetase from methicillin resistant Staphylococcus aureus (MRSA).

NAD synthetase (NadE) catalyzes the last step in NAD biosynthesis, transforming deamido-NAD+ into NAD+ by a two-step reaction with co-substrates ATP and amide donor ammonia. In this study, we report the crystal structure of Staphylococcus aureus NAD synthetase enzyme (saNadE) at 2.3 Å resolution. We used this structure to perform molecular dynamics simulations of apo-enzyme, enzyme-substrate (NadE with ATP and NaAD) and enzyme-intermediate complexes (NadE with NaAD-AMP) to investigate key binding interactions and explore the conformational transitions and flexibility of the binding pocket. Our results show large shift of N-terminal region in substrate bound form which is important for ATP binding. Substrates drive the correlated movement of loop regions surrounding it as well as some regions distal to the active site and stabilize them at complex state. Principal component analysis of atomic projections distinguish feasible trajectories to delineate distinct motions in enzyme-substrate to enzyme-intermediate states. Our results suggest mixed binding involving dominant induced fit and conformational selection. MD simulation extracted ensembles of NadE could potentially be utilized for in silico screening and structure based design of more effective Methicillin Resistant Staphylococcus aureus (MRSA) inhibitors.