In silico prediction of mutant HIV-1 proteases cleaving a target sequence

HIV-1 protease represents an appealing system for directed enzyme re-design, since it has various different endogenous targets, a relatively simple structure and it is well studied. Recently Chaudhury and Gray (Structure (2009) 17: 1636 -- 1648) published a computational algorithm to discern the specificity determining residues of HIV-1 protease. In this paper we present two computational tools aimed at re-designing HIV-1 protease, derived from the algorithm of Chaudhuri and Gray. First, we present an energy-only based methodology to discriminate cleavable and non cleavable peptides for HIV-1 proteases, both wild type and mutant. Secondly, we show an algorithm we developed to predict mutant HIV-1 proteases capable of cleaving a new target substrate peptide, different from the natural targets of HIV-1 protease. The obtained in silico mutant enzymes were analyzed in terms of cleavability and specificity towards the target peptide using the energy-only methodology. We found two mutant proteases as best candidates for specificity and cleavability towards the target sequence

A Computational Methodology to Screen Activities of Enzyme Variants

We present a fast computational method to efficiently screen enzyme activity. In the presented method, the effect of mutations on the barrier height of an enzyme-catalysed reaction can be computed within 24 hours on roughly 10 processors. The methodology is based on the PM6 and MOZYME methods as implemented in MOPAC2009, and is tested on the first step of the amide hydrolysis reaction catalyzed by Candida Antarctica lipase B (CalB) enzyme. The barrier heights are estimated using adiabatic mapping and are shown to give barrier heights to within 3kcal/mol of B3LYP/6-31G(d)//RHF/3-21G results for a small model system. Relatively strict convergence criteria (0.5kcal/(mol{\AA})), long NDDO cutoff distances within the MOZYME method (15{\AA}) and single point evaluations using conventional PM6 are needed for reliable results. The generation of mutant structure and subsequent setup of the semiempirical calculations are automated so that the effect on barrier heights can be estimated for hundreds of mutants in a matter of weeks using high performance computing

Polygons on a Rotating Fluid Surface

We report a novel and spectacular instability of a fluid surface in a rotating system. In a flow driven by rotating the bottom plate of a partially filled, stationary cylindrical container, the shape of the free surface can spontaneously break the axial symmetry and assume the form of a polygon rotating rigidly with a speed different from that of the plate. With water we have observed polygons with up to 6 corners. It has been known for many years that such flows are prone to symmetry breaking, but apparently the polygonal surface shapes have never been observed. The creation of rotating internal waves in a similar setup was observed for much lower rotation rates, where the free surface remains essentially flat. We speculate that the instability is caused by the strong azimuthal shear due to the stationary walls and that it is triggered by minute wobbling of the rotating plate. The slight asymmetry induces a tendency for mode-locking between the plate and the polygon, where the polygon rotates by one corner for each complete rotation of the plate

Chemical control of bacterial epibiosis on ascidians

Two co-occurring ascidians of the Pacific subtidal, Polyclinum planum and Cystodytes lobatus, were found to exhibit remarkably different numbers of surface bacteria. On average, epibacterial densities on P. planum were 140 times greater than those on C lobatus as measured by plate-counting methods. Organic extracts of both ascidian species were tested for antimicrobial activities and effects on bacterial settlement. Bacterial settlement was measured using a new bioassay described in this paper. The results of the settlement bioassay clearly demonstrate that extracts of the little-colonized C. lobatus deter bacterial settlement while extracts of the heavily colonized P. planum induce settlement relative to the control. In addition to reducing bacterial settlement, extracts of C. lobatus colonies exhibited varying, but generally low, levels of antiraicrobial activity against, on average, one-half of the 36 strains of marine bacteria tested. On an interspecific level, including 11 ascidian species screened in a pilot study, fouling-deterring activities were correlated with epibacterial abundances while antimicrobial activity was not. It is concluded that the chemical control of bacterial settlement, possibly complemented by antimicrobial toxicity, provides an accurate model to explain the dramatically different bacterial abundance on the surfaces of the ascidian species studied. This investigation presents evidence that non-toxic metabolites influence bacterial settlement and, in this way, may function to regulate bacterial epibiosis on the surfaces of some marine invertebrates

Statistics of lattice animals (polyominoes) and polygons

We have developed an improved algorithm that allows us to enumerate the number of site animals (polyominoes) on the square lattice up to size 46. Analysis of the resulting series yields an improved estimate, $\tau = 4.062570(8)$, for the growth constant of lattice animals and confirms to a very high degree of certainty that the generating function has a logarithmic divergence. We prove the bound $\tau > 3.90318.$ We also calculate the radius of gyration of both lattice animals and polygons enumerated by area. The analysis of the radius of gyration series yields the estimate $\nu = 0.64115(5)$, for both animals and polygons enumerated by area. The mean perimeter of polygons of area $n$ is also calculated. A number of new amplitude estimates are given.Comment: 10 pages, 2 eps figure

Mobile bag technique for estimation of nutrient digestibility when hay is supplemented with alternative fibrous feedstuffs in horses

publishedVersio

Study of the Potts Model on the Honeycomb and Triangular Lattices: Low-Temperature Series and Partition Function Zeros

We present and analyze low-temperature series and complex-temperature partition function zeros for the $q$-state Potts model with $q=4$ on the honeycomb lattice and $q=3,4$ on the triangular lattice. A discussion is given as to how the locations of the singularities obtained from the series analysis correlate with the complex-temperature phase boundary. Extending our earlier work, we include a similar discussion for the Potts model with $q=3$ on the honeycomb lattice and with $q=3,4$ on the kagom\'e lattice.Comment: 33 pages, Latex, 9 encapsulated postscript figures, J. Phys. A, in pres

Linear response functions for a vibrational configuration interaction state

Linear response functions are implemented for a vibrational configuration interaction state allowing accurate analytical calculations of pure vibrational contributions to dynamical polarizabilities. Sample calculations are presented for the pure vibrational contributions to the polarizabilities of water and formaldehyde. We discuss the convergence of the results with respect to various details of the vibrational wave function description as well as the potential and property surfaces. We also analyze the frequency dependence of the linear response function and the effect of accounting phenomenologically for the finite lifetime of the excited vibrational states. Finally, we compare the analytical response approach to a sum-over-states approac

BioFET-SIM Web Interface: Implementation and Two Applications

We present a web interface for the BioFET-SIM program. The web interface allows to conveniently setup calculations based on the BioFET-SIM multiple charges model. As an illustration, two case studies are presented. In the first case, a generic peptide with opposite charges on both ends is inverted in orientation on a semiconducting nanowire surface leading to a corresponding change in sign of the computed sensitivity of the device. In the second case, the binding of an antibody/antigen complex on the nanowire surface is studied in terms of orientation and analyte/nanowire surface distance. We demonstrate how the BioFET-SIM web interface can aid in the understanding of experimental data and postulate alternative ways of antibody/antigen orientation on the nanowire surface

A computational method for the systematic screening of reaction barriers in enzymes:searching for Bacillus circulans xylanase mutants with greater activity towards a synthetic substrate

We present a semi-empirical (PM6-based) computational method for systematically estimating the effect of all possible single mutants, within a certain radius of the active site, on the barrier height of an enzymatic reaction. The intent of this method is not a quantitative prediction of the barrier heights, but rather to identify promising mutants for further computational or experimental study. The method is applied to identify promising single and double mutants of Bacillus circulans xylanase (BCX) with increased hydrolytic activity for the artificial substrate ortho-nitrophenyl β-xylobioside (ONPX2). The estimated reaction barrier for wild-type (WT) BCX is 18.5 kcal/mol, which is in good agreement with the experimental activation free energy value of 17.0 kcal/mol extracted from the observed kcat using transition state theory (Joshi et al., 2001). The PM6 reaction profiles for eight single point mutations are recomputed using FMO-MP2/PCM/6-31G(d) single points. PM6 predicts an increase in barrier height for all eight mutants while FMO predicts an increase for six of the eight mutants. Both methods predict that the largest change in barrier occurs for N35F, where PM6 and FMO predict a 9.0 and 15.8 kcal/mol increase, respectively. We thus conclude that PM6 is sufficiently accurate to identify promising mutants for further study. We prepared a set of all theoretically possible (342) single mutants in which every amino acid of the active site (except for the catalytically active residues E78 and E172) was mutated to every other amino acid. Based on results from the single mutants we construct a set of 111 double mutants consisting of all possible pairs of single mutants with the lowest barrier for a particular position and compute their reaction profile. None of the mutants have, to our knowledge, been prepared experimentally and therefore present experimentally testable predictions