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ABSTRACTA wetland is an ecosystem, which is inundated by water. Several biogeochemical processes related to wetlands place it into an important environmental system class. This study employs for the first time, a data-driven methodology, specifically one based on system identification, to develop a nonlinear-autoregressive-exogenous-input model structure corresponding to the tropical wetland dynamics, as an alternative to the classical topographic modelling approaches. Altogether 94 wetland sites from Amazon, Africa, and Asia constitute the model structure development. The topographical variations among the wetlands entail a unique set of model parameters for each wetland site, which is achieved by using the approximate Bayesian computation sequential Monte Carlo method. The performance of the single model structure is subsequently evaluated by simulating all the sample wetland sites. The obtained wetland model structure in this study can be used to parameterise any tropical wetland site as well as simulate their dynamical characteristics.

Primary productivity is enhanced within a few kilometres of icebergs in the Weddell Sea owing to the input of terrigeneous nutrients and trace elements during iceberg melting. However, the influence of giant icebergs, over 18 km in length, on marine primary production in the Southern Ocean is less well studied. Here we present an analysis of 175 satellite images of open ocean colour before and after the passage of 17 giant icebergs between 2003 and 2013. We detect substantially enhanced chlorophyll levels, typically over a radius of at least 4–10 times the iceberg’s length, that can persist for more than a month following passage of a giant iceberg. This area of influence is more than an order of magnitude larger than that found for sub-kilometre scale icebergs or in ship-based surveys of giant icebergs. Assuming that carbon export increases by a factor of 5–10 over the area of influence, we estimate that up to a fifth of the Southern Ocean’s downward carbon flux originates with giant iceberg fertilization. We suggest that, if giant iceberg calving increases this century as expected, this negative feedback on the carbon cycle may become more important.

In this work we measured 1H NMR chemical shifts for the ribonuclease barnase at pressures from 3 MPa to 200 MPa, both free and bound to d(CGAC). Shift changes with pressure were used as restraints to determine the change in structure with pressure. Free barnase is compressed by ∼0.7%. The largest changes are on the ligand-binding face close to Lys-27, which is the recognition site for the cleaved phosphate bond. This part of the protein also contains the buried water molecules. In the presence of d(CGAC), the compressibility is reduced by ∼70% and the region of structural change is altered: the ligand-binding face is now almost incompressible, whereas changes occur at the opposite face. Because compressibility is proportional to mean square volume fluctuation, we conclude that in free barnase, volume fluctuation is largest close to the active site, but when the inhibitor is bound, the fluctuations become much smaller and are located mainly on the opposite face. The timescale of the fluctuations is nanoseconds to microseconds, consistent with the degree of ordering required for the fluctuations, which are intermediate between rapid uncorrelated side-chain dynamics and slow conformational transitions. The high-pressure technique is therefore useful for characterizing motions on this relatively inaccessible timescale.

Binary kernel discrimination (BKD) uses a training set of compounds, for which structural and qualitative activity data are available, to produce a model that can then be applied to the structures of other compounds in order to predict their likely activity. Experiments with the MDL Drug Data Report database show that the optimal value of the smoothing parameter, and hence the predictive power of BKD, is crucially dependent on the number of false positives in the training set. It is also shown that the best results for BKD are achieved using one particular optimization method for the determination of the smoothing parameter that lies at the heart of the method and using the Jaccard/Tanimoto coefficient in the kernel function that is used to compute the similarity between a test set molecule and the members of the training set.

This paper discusses the use of binary kernel discrimination (BKD) for identifying potential active compounds in lead-discovery programs. BKD was compared with established virtual screening methods in a series of experiments using pesticide data from the Syngenta corporate database. It was found to be superior to methods based on similarity searching and substructural analysis but inferior to a support vector machine. Similar conclusions resulted from application of the methods to a pesticide data set for which categorical activity data were available.

Similarity searching using a single bioactive reference structure is a well-established technique for accessing chemical structure databases. This paper describes two extensions of the basic approach. First, we discuss the use of group fusion to combine the results of similarity searches when multiple reference structures are available. We demonstrate that this technique is notably more effective than conventional similarity searching in scaffold-hopping searches for structurally diverse sets of active molecules; conversely, the technique will do little to improve the search performance if the actives are structurally homogeneous. Second, we make the assumption that the nearest neighbors resulting from a similarity search, using a single bioactive reference structure, are also active and use this assumption to implement approximate forms of group fusion, substructural analysis, and binary kernel discrimination. This approach, called turbo similarity searching, is notably more effective than conventional similarity searching.

Fingerprint-based similarity searching is widely used for virtual screening when only a single bioactive reference structure is available. This paper reviews three distinct ways of carrying out such searches when multiple bioactive reference structures are available:  merging the individual fingerprints into a single combined fingerprint; applying data fusion to the similarity rankings resulting from individual similarity searches; and approximations to substructural analysis. Extended searches on the MDL Drug Data Reportdatabase suggest that fusing similarity scores is the most effective general approach, with the best individual results coming from the binary kernel discrimination technique.

This paper discusses the use of several rank-based virtual screening methods for prioritizing compounds in lead-discovery programs, given a training set for which both structural and bioactivity data are available. Structures from the NCI AIDS data set and from the Syngenta corporate database were represented by two types of fragment bit-string and by sets of high-level molecular features. These representations were processed using binary kernel discrimination, similarity searching, substructural analysis, support vector machine, and trend vector analysis, with the effectiveness of the methods being judged by the extent to which active test set molecules were clustered toward the top of the resultant rankings. The binary kernel discrimination approach yielded consistently superior rankings and would appear to have considerable potential for chemical screening applications.

A chemical hyperstructure is a single graph representation of a set of molecules that minimizes the degree of structural redundancy in the data set. This paper describes the use of a genetic algorithm to generate an activity-weighted chemical hyperstructure (AWCH) by sequentially mapping each molecule in the data set to the hyperstructure and then assigning activity and inactivity frequency weights to the nodes and edges of the hyperstructure. Experiments with several data sets demonstrate the level of activity clustering in an AWCH.

Liver fatty acid-binding protein (FABP) is able to bind to anionic phospholipid vesicles under conditions of low ionic strength. This binding results in the release of ligand, the fluorescent fatty acid analogue 11-dansylaminoundecanoic acid (DAUDA), with loss of fluorescence intensity (Davies, J. K., Thumser, A. E. A., and Wilton, D. C. (1999) Biochemistry 38, 16932-16940). Using a strategy of charge reversal mutagenesis, the potential role of specific cationic residues in promoting interfacial binding of FABP to anionic phospholipid vesicles has been investigated. Cationic residues chosen included those within the alpha-helical region (Lys-20, Lys-31, and Lys-33) and those that make a significant contribution to the positive surface potential of the protein (Lys-31, Lys-36, Lys-47, Lys-57, and Arg-126). Only three cationic residues make a significant contribution to interfacial binding, and these residues (Lys-31, Lys-36, and Lys-57) are all located within the ligand portal region, where the protein may be predicted to exhibit maximum disorder. The binding of tryptophan mutants, F3W, F18W, and C69W, to dioleoylphosphatidylglycerol vesicles, containing 5 mol% of the fluorescent phospholipid dansyldihexadecanoylphosphatidylethanolamine, was monitored by fluorescence resonance energy transfer (FRET). All three mutants showed enhanced dansyl fluorescence due to FRET on addition of phospholipid to protein; however, this fluorescence was considerably greater with the F3W mutant, consistent with the N-terminal region of the protein coming in close proximity to the phospholipid interface. These results were confirmed by succinimide quenching studies. Overall, the results indicate that the portal region of liver FABP and specifically Lys-31, Lys-36, and Lys-57 are involved in the interaction with the interface of anionic vesicles and that the N-terminal region of the protein undergoes a conformational change, resulting in DAUDA release.

Rigid ring systems can be used to position receptor-binding functional groups in 3D space and they thus play an increasingly important role in the design of combinatorial libraries. This paper discusses the use of shape-similarity methods to identify ring systems that are structurally similar to, and aligned with, a user-defined target ring system. These systems can be used as alternative scaffolds for the construction of a combinatorial library.

Liver fatty acid binding protein (LFABP) is unique among the various types of FABPs in that it can bind a variety of ligands in addition to fatty acids. LFABP is able to bind long chain fatty acids with a 2:1 stoichiometry and the crystal structure has identified two fatty acid binding sites in the binding cavity. The presumed primary site (site 1) involves the fatty acid binding with the carboxylate group buried in the cavity whereas the fatty acid at site 2 has the carboxylate group solvent-exposed within the ligand portal region and in the vicinity of α-helix II. The α-helical region contains three cationic residues, K20, K31, K33 and modelling studies suggest that K31 on α-helix II could make an electrostatic contribution to anionic ligands binding to site 2. The preparation of three charge reversal mutants of LFABP, K20E, K31E and K33E has allowed an investigation of the role of site 2 in ligand binding, particularly those ligands with a bulky anionic head group. The binding of oleoyl CoA, lysophosphatidic acid, lysophosphatidylcholine, lithocholic acid and taurolithocholate 3-sulphate to LFABP has been studied using the α-helical mutants. The results support the concept that such ligands bind at site 2 of LFABP where solvent exposure allows the accommodation of their bulky anionic group.

The antibacterial properties of human group IIA secreted phospholipase A2against Gram-positive bacteria as a result of membrane hydrolysis have been reported. Using Micrococcus luteusas a model system, we demonstrate the very high specificity of this human enzyme for such hydrolysis compared with the group IB, IIE, IIF, V, and X human secreted phospholipase A2s. A unique feature of the group IIA enzyme is its very high pI due to a large excess of cationic residues on the enzyme surface. The importance of this global positive charge in bacterial cell membrane hydrolysis and bacterial killing has been examined using charge reversal mutagenesis. The global positive charge on the enzyme surface allows penetration through the bacterial cell wall, thus allowing access of this enzyme to the cell membrane. Reduced bacterial killing was associated with the loss of positive charge and reduced cell membrane hydrolysis. All mutants were highly effective in hydrolyzing the bacterial membrane of cells in which the cell wall was permeabilized with lysozyme. These same overall characteristics were also seen with suspensions of Staphylococcus aureusand Listeria innocua, where cell membrane hydrolysis and antibacterial activity of human group IIA enzyme was also lost as a result of charge reversal mutagenesis.

We present the theory of the electron propagator perturbed by an external electric field and show how it can be used to calculate a variety of one-electron linear response properties that are accurate through second order in electron correlation. Some illustrative calculations are discussed. © 2000 John Wiley & Sons, Inc. Int J Quant Chem 77: 625–640, 2000

Hepatic-FABP was labelled at cysteine-69 with the fluorescent environmentally sensitive reporter group AEDANS. The labelled protein had an emission maximum at 465 nm indicating that cysteine-69 was buried in a non-polar environment. The modified protein was still able to bind ligands such as oleic acid, oleoyl CoA and haem. The affinity of AEDANS-FABP for haem was unaltered as compared with the native protein indicating that cysteine-69 must be remote from the ligand binding site. The binding of oleic acid did not significantly perturb the fluorescence emission spectrum of the fluorescent reporter group suggesting that there are not large conformational changes in the region of cysteine-69 on fatty acid binding. The binding of stoichiometric amounts of oleoyl CoA was accompanied by a small fluorescence enhancement which suggests that fatty acyl CoAs may interact with other regions of the FABP molecule not involved in fatty acid binding.

Differences between real tourism satellite accounts and the WTTC/WEFA ‘simulated tourism satellite accounts’ are described. These differences are both conceptual and methodological, including the fact that the World Tourism and Travel Council (WTTC) methodology fails to conform to the World Tourism Organization/UN definitions. Some empirical problems associated with the WTTC methodology also are detailed. These include the inappropriate use of Consumer Price Index weightings and WTTC's reliance on US data sources to make inferences about other nations' economies. The paper concludes that the WTTC methodology does not represent a ‘simulated tourism satellite account’.

Tetrahymanol biosynthesis by the protozoan Tetrahymena pyriformis was progressively inhibited by the inclusion of cholesterol in the growth medium. Studies with labelled precursors of tetrahymanol have established that there are two major sites of inhibition in whole cells. The inhibition at the first site, between acetate and mevalonate, occurred rapidly after addition of cholesterol. The activity of 3-hydroxy-3-methylglutaryl-CoA reductase (EC 1.1.1.34), a predominantly cytosolic enzyme in this organism, was not inhibited in cholesterol-grown cells nor by addition of cholesterol directly to the assay medium. The second major site of inhibition in whole cells is between mevalonate and squalene and this is accompanied by inhibition of the enzyme that converts farnesyl-pyrophosphate into squalene (squalene synthetase). Squalene cyclase is partially inhibited. The conversion of mevalonate into tetrahymanol in vitro was not inhibited by the addition of cholesterol to the assay medium. Tetrahymanol added to the culture medium is taken up by the cells but does not inhibit endogenous biosynthesis. It is suggested that cholesterol inhibits the later stages of tetrahymanol biosynthesis by causing a change in membrane structure and function which alters the activity of membrane-bound enzymes.

The identification of a database subset in which all pairs of molecules have similarities less than some user-defined threshold is shown to be equivalent to the graph-theoretic problem of clique-detection. A comparison of several different clique-detection algorithms demonstrates that the algorithm due to Babel is by far the most efficient for this particular problem. The database subsets identified by the Babel algorithm are slightly more diverse than those identified by alternative, less time-consuming algorithms for dissimilarity-based compound selection.

The breakdown of normal substrates by lipases requires an interfacial binding step prior to hydrolysis. Interfacial binding and subsequent hydrolysis will be affected by the lipid components and hence physical properties of the substrate surface. In order to investigate in detail the effect of lipid structure on the activity of lipoprotein lipase (LPL), triolein-containing emulsion particles of defined composition have been used as substrates. In addition, lipase activity has been measured using a continuous fluorescence displacement assay that monitors the release of long-chain fatty acids as an alternative to normal radiochemical assays. Using this fluorescence assay, rates of hydrolysis of triolein were the same as when using a standard radiochemical assay under identical conditions. Activation by apolipoprotein CII was very similar by both methods; however, the extent of activation (2Ő3-fold) was less than has been reported previously using different assay conditions. In order to investigate the effect of cholesterol on LPL activity, emulsion particles were prepared in which the cholesterol/egg-phosphatidylcholine ratio was increased up to a 1:1 molar ratio. A pronounced stimulatory effect of cholesterol was observed under these assay conditions, with up to a 5-fold increase in rate compared with emulsion particles without cholesterol. Since high molar ratios of cholesterol are reported to exclude triacylglycerol from the phospholipid surface [Spooner and Small (1987) Biochemistry 26, 5820Ő5825], these results are not consistent with a mechanism involving LPL hydrolysis of surface triacylglycerol. Instead, they support an interfacial penetration model, allowing the enzyme's active site direct access to triacylglycerol in the lipoprotein core. Perturbation of the surface phospholipid monolayer of the emulsion particle as a result of hydrolysis by Naja naja phospholipase A2 resulted in a 10-fold activation of LPL, providing further support for an interfacial penetration model. The stimulatory effect of apolipoprotein CII was not modulated by modification of the interface with cholesterol.

The physiological role of liver fatty acid-binding protein (L-FABP) has yet to be clarified. An important feature of this member of the family of intracellular lipid-binding proteins is the wide range of compounds that have been identified as potential physiological ligands. By using recombinant L-FABP, the binding of cholesterol, bile salts and their derivatives has been investigated under conditions that allow a direct comparison of the binding affinities of these ligands for fatty acids. The results demonstrate an inability of L-FABP to bind cholesterol, although the anionic derivative, cholesteryl sulphate, will bind under similar assay conditions. Of the bile salts examined, lithocholate and taurolithocholate sulphate showed the greatest binding to L-FABP. It is proposed that an important function of L-FABP is to bind certain physiological amphipathic anions, thus preventing the ‘free’ concentrations of these compounds from exceeding their critical micelle concentration, which could result in cell damage.

Rat liver fatty acid-binding protein (FABP) is able to accommodate a wide range of non-polar anions in addition to long-chain fatty acids. This property means that the liver protein is functionally different from other FABPs from intestine, muscle and adipose tissue that have a more restricted ligand specificity and stoichiometry. The availability of crystal structures for the latter proteins has highlighted the importance of two arginine residues that are involved in the binding of the fatty acid carboxylate. Only one of these arginine residues, arginine-122, is conserved in liver FABP, whereas the other arginine, at position 102, is replaced by a threonine. In order to gain further insight into the nature of ligand interactions with liver FABP these key residues (102 and 122) have been changed by site-directed mutagenesis. The results with an R122Q mutant highlight the critical role of this arginine in determining ligand affinity, while similar but less dramatic effects were observed with the T102Q mutant. The double mutant T102Q/R122Q was expressed but had lost the ability to bind fluorescent ligands. It is concluded that Arg-122 plays a role in accommodating the carboxylate group of at least one fatty acid. It is proposed that physiological ligands with more bulky headgroups, such as lysophospholipids, acyl-CoA and mono-olein, bind with the headgroups in a solvent-exposed location near the portal region of the protein. The portal region is suggested to be more flexible in the mutants (R122Q and to a lesser extent T102Q). The net result is that the ligand specificity of the R122Q mutant changes to that of a protein with enhanced affinity for acyl-CoA, lysophospholipids and mono-olein.

The purpose of this paper is to clarify the question of nonexistence of solutions for the superposition method in acoustic radiation and scattering problems. Recently a number of authors have implemented this method and have claimed that nonexistence or nonuniqueness difficulties do not arise. Numerical evidence appears to support this view. However, this paper shows that the superposition method is a discrete approximation to an equation that does suffer from nonexistence of its solution at certain frequencies. Nonexistence of solutions is demonstrated with some numerical examples and an improved method is discussed.

The identification of a database subset in which all pairs of molecules have similarities less than some user-defined threshold is shown to be equivalent to the graph-theoretic problem of clique-detection. A comparison of several different clique-detection algorithms demonstrates that the algorithm due to Babel is by far the most efficient for this particular problem. The database subsets identified by the Babel algorithm are slightly more diverse than those identified by alternative, less time-consuming algorithms for dissimilarity-based compound selection.

The identification of a database subset in which all pairs of molecules have similarities less than some user-defined threshold is shown to be equivalent to the graph-theoretic problem of clique-detection. A comparison of several different clique-detection algorithms demonstrates that the algorithm due to Babel is by far the most efficient for this particular problem. The database subsets identified by the Babel algorithm are slightly more diverse than those identified by alternative, less time-consuming algorithms for dissimilarity-based compound selection.

Human group IIa phospholipase A2(hIIa-PLA2) is a highly basic protein that is secreted from a number of cells during inflammation and may play a role in arachidonate liberation and in destruction of invading bacteria. It has been proposed that rodent group IIa PLA2is anchored to cell surfaces via attachment to heparan sulfate proteoglycan and that this interaction facilitates lipolysis. hIIa-PLA2 contains 13 lysines, 2 histidines, and 10 arginines that fall into 10 clusters. A panel of 26 hIIa-PLA2 mutants were prepared in which 1–4 basic residues in each cluster were changed to glutamate or aspartate (charge reversal). A detailed analysis of the affinities of these mutants for anionic vesicles and for heparin and heparan sulfate in vitroand of the specific activities of these proteins for hydrolysis of vesiclesin vitroand of living cell membranes reveal the following trends: 1) the affinity of hIIa-PLA2 for heparin and heparan sulfate is modulated not by a highly localized site of basic residues but by diffuse sites that partially overlap with the interfacial binding site. In contrast, only those residues on the interfacial binding site of hIIa-PLA2 are involved in binding to membranes; 2) the relative ability of these mutants to hydrolyze cellular phospholipids when enzymes were added exogenously to CHO-K1, NIH-3T3, and RAW 264.7 cells correlates with their relative in vitroaffinity for vesicles and not with their affinity for heparin and heparan sulfate. 3) The rates of exogenous hIIa-PLA2-catalyzed fatty acid release from wild type CHO-K1 cells and two mutant lines, one lacking glycosaminoglycan and one lacking heparan sulfate, were similar. Thus basic residues that modulate interfacial binding are important for plasma membrane fatty acid release by exogenously added hIIa-PLA2. Binding of hIIa-PLA2 to cell surface heparan sulfate does not modulate plasma membrane phospholipid hydrolysis by exogenously added hIIa-PLA2.

The less polar fluorescent analogue of ATP, 2′(3′)-trinitrophenyl-5′-triphosphate bound to rat liver fatty acid-binding protein with high affinity ( Kd6.3 × 10 −6M) and 1:1 molar stoichiometry. This probe bound to the fatty acid binding site of the protein and was displaced by oleic acid and oleoyl CoA. High concentrations of ATP did not cause significant displacement of the fluorescent ATP analogue. Since the anionic part of this molecule is the triphosphate group it is difficult to envisage this group being accommodated at an anion binding site within the non-polar core of this protein as is the case with other fatty acid binding proteins. Therefore it is anticipated that the ligand must bind to liver fatty acid-binding protein with these triphosphate group surface exposed. Caution must be exercised when using the more hydrophobic fluorescent analogue of ATP to investigate the ATP binding properties of proteins.

The chemical synthesis and metabolism of [3α-3H]cholesta-5,7,9(11)-trien-3β-ol is described. This radioactive fluorescent sterol probe cannot be rearranged or converted to cholesterol by rat liver but is readily esterified.

The less polar fluorescent analogue of ATP, 2′(3′)‐trinitrophenyl‐5′‐triphosphate bound to rat liver fatty acid‐binding protein with high affinity (Kd6.3 × 10−6M) and 1:1 molar stoichiometry. This probe bound to the fatty acid binding site of the protein and was displaced by oleic acid and oleoyl CoA. High concentrations of ATP did not cause significant displacement of the fluorescent ATP analogue. Since the anionic part of this molecule is the triphosphate group it is difficult to envisage this group being accommodated at an anion binding site within the non‐polar core of this protein as is the case with other fatty acid binding proteins. Therefore it is anticipated that the ligand must bind to liver fatty acid‐binding protein with these triphosphate group surface exposed. Caution must be exercised when using the more hydrophobic fluorescent analogue of ATP to investigate the ATP binding properties of proteins.

This paper proposes and demonstrates the accuracy of new approximations to the spectral response functions of single poles with arbitrary pole potential distributions, in the presence of an underlayer. In particular, two approximations to the spectral response for the conventional constant potential single pole are compared.