The Adaptive Poisson-Boltzmann Solver (APBS) software was developed to solve the equations of continuum electrostatics for large biomolecular assemblages that have provided impact in the study of a broad range of chemical, biological, and biomedical applications. APBS addresses the three key technology challenges for understanding solvation and electrostatics in biomedical applications: accurate and efficient models for biomolecular solvation and electrostatics, robust and scalable software for applying those theories to biomolecular systems, and mechanisms for sharing and analyzing biomolecular electrostatics data in the scientific community. To address new research applications and advancing computational capabilities, we have continually updated APBS and its suite of accompanying software since its release in 2001. In this article, we discuss the models and capabilities that have recently been implemented within the APBS software package including a Poisson-Boltzmann analytical and a semi-analytical solver, an optimized boundary element solver, a geometry-based geometric flow solvation model, a graph theory-based algorithm for determining pK values, and an improved web-based visualization tool for viewing electrostatics.

We provide an overview of the key architectural features of recent microprocessor designs and describe the programming model and abstractions provided by OpenCL, a new parallel programming standard targeting these architectures.

Cohesin is crucial for proper chromosome segregation but also regulates gene transcription and organism development by poorly understood mechanisms. Using genome-wide assays in Drosophila developing wings and cultured cells, we find that cohesin functionally interacts with Polycomb group (PcG) silencing proteins at both silenced and active genes. Cohesin unexpectedly facilitates binding of Polycomb Repressive Complex 1 (PRC1) to many active genes, but their binding is mutually antagonistic at silenced genes. PRC1 depletion decreases phosphorylated RNA polymerase II and mRNA at many active genes but increases them at silenced genes. Depletion of cohesin reduces long-range interactions between Polycomb Response Elements in the invected-engrailed gene complex where it represses transcription. These studies reveal a previously unrecognized role for PRC1 in facilitating productive gene transcription and provide new insights into how cohesin and PRC1 control development.

Cohesin is a well-known mediator of sister chromatid cohesion, but it also influences gene expression and development. These non-canonical roles of cohesin are not well understood, but are vital: gene expression and development are altered by modest changes in cohesin function that do not disrupt chromatid cohesion. To clarify cohesin's roles in transcription, we measured how cohesin controls RNA polymerase II (Pol II) activity by genome-wide chromatin immunoprecipitation and precision global run-on sequencing. On average, cohesin-binding genes have more transcriptionally active Pol II and promoter-proximal Pol II pausing than non-binding genes, and are more efficient, producing higher steady state levels of mRNA per transcribing Pol II complex. Cohesin depletion frequently decreases gene body transcription but increases pausing at cohesin-binding genes, indicating that cohesin often facilitates transition of paused Pol II to elongation. In many cases, this likely reflects a role for cohesin in transcriptional enhancer function. Strikingly, more than 95% of predicted extragenic enhancers bind cohesin, and cohesin depletion can reduce their association with Pol II, indicating that cohesin facilitates enhancer-promoter contact. Cohesin depletion decreases the levels of transcriptionally engaged Pol II at the promoters of most genes that don't bind cohesin, suggesting that cohesin controls expression of one or more broadly acting general transcription factors. The multiple transcriptional roles of cohesin revealed by these studies likely underlie the growth and developmental deficits caused by minor changes in cohesin activity.

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The use of Mn 2+ as the divalent cation cofactor in polymerase-catalyzed reactions instead of Mg 2+ often diminishes the stringency of substrate selection and incorporation fidelity. We have solved the complete kinetic mechanism for single nucleotide incorporation catalyzed by the RNA-dependent RNA polymerase from poliovirus (3D pol ) in the presence of Mn 2+ . The steps employed during a single cycle of nucleotide incorporation are identical to those employed in the presence of Mg 2+ and include a conformational-change step after nucleotide binding to achieve catalytic competence of the polymerase-primer/template-nucleotide complex. In the presence of Mn 2+ , the conformationalchange step is the primary determinant of enzyme specificity, phosphoryl transfer appears as the sole rate-limiting step for nucleotide incorporation, and the rate of phosphoryl transfer is the same for all nucleotides: correct and incorrect. Because phosphoryl transfer is the rate-limiting step in the presence of Mn 2+ , it was possible to determine that the maximal phosphorothioate effect in this system is in the range of 8-11. This information permitted further interrogation of the nucleotideselection process in the presence of Mg 2+ , highlighting the capacity of this cation to permit the enzyme to use the phosphoryl-transfer step for nucleotide selection. The inability of Mn 2+ to support a reduction in the efficiency of phosphoryl transfer when incorrect substrates are employed is the primary explanation for the loss of fidelity observed in the presence of this cofactor. We propose that the conformational change involves reorientation of the triphosphate moiety of the bound nucleotide into a conformation that permits binding of the second metal ion required for catalysis. In the presence of Mg 2+ , this conformation requires interactions with the enzyme that permit a reduction in catalytic efficiency to occur during an attempt to incorporate an incorrect nucleotide. Adventitious interactions in the cofactor-binding site with bound Mn 2+ may diminish fidelity by compensating for interaction losses used to modulate catalytic efficiency when incorrect nucleotides are bound in the presence of Mg 2+ .All nucleic acid polymerases require two divalent cations as cofactors to catalyze phosphoryl transfer (1). The first metal ion is brought into the active site complexed to the triphosphate moiety of the nucleotide substrate. This metal ion may facilitate formation of the conformation of the triphosphate required for nucleophilic attack of the α-phosphorus atom. The second metal ion is required to lower the pK a of the primer 3′-OH to facilitate formation of the nucleophile required for catalysis. Mg 2+ is thought to be the divalent cation employed by most polymerases known (2); however, activity is usually supported by other divalent cations as well (3-7). In a Δ This work was supported, in part, by a Howard Temin Award (CA75118) from the NCI, National Institutes of Health, and by a grant (AI45818) from the NIAID, National Institute...

The prevailing "plug-in-the-bottle" model suggests that macrolide antibiotics inhibit translation by binding inside the ribosome tunnel and indiscriminately arresting the elongation of every nascent polypeptide after the synthesis of six to eight amino acids. To test this model, we performed a genome-wide analysis of translation in azithromycin-treated Staphylococcus aureus. In contrast to earlier predictions, we found that the macrolide does not preferentially induce ribosome stalling near the 5′ end of mRNAs, but rather acts at specific stalling sites that are scattered throughout the entire coding region. These sites are highly enriched in prolines and charged residues and are strikingly similar to other ligandindependent ribosome stalling motifs. Interestingly, the addition of structurally related macrolides had dramatically different effects on stalling efficiency. Our data suggest that ribosome stalling can occur at a surprisingly large number of low-complexity motifs in a fashion that depends only on a few arrest-inducing residues and the presence of a small molecule inducer.antibiotic | ribosome stalling | Staphylococcus aureus

For over four decades, two competing mechanisms of ligand recognition – conformational selection and induced-fit - have dominated our interpretation of protein allostery. Defining the mechanism broadens our understanding of the system and impacts our ability to design effective drugs and new therapeutics. Recent kinetics studies demonstrate that trypsin-like proteases exist in equilibrium between two forms: one fully accessible to substrate (E) and the other with the active site occluded (E*). Analysis of the structural database confirms existence of the E* and E forms and vouches for the allosteric nature of the trypsin fold. Allostery in terms of conformational selection establishes an important paradigm in the protease field and enables protein engineers to expand the repertoire of proteases as therapeutics.