FINDSITELHM: A Threading-Based Approach to Ligand Homology
                    Modeling
[['Ligand virtual screen is widely used in rational drug discovery [1] , [2] .', 'The first stage of', 'structure-based ligand screening is the prediction of the binding mode adopted by', 'the small molecule complexed to its target receptor protein; a variety of algorithms', 'have been developed to achieve this goal [3] , [4] .', 'The next step is to', 'estimate the relative binding affinity of the docked ligands [5] , [6] .', 'Of course , it is not', 'sufficient that a given ligand binds favorably to given protein; rather , to minimize', 'side effects , it must also bind selectively .', 'Classical molecular docking has been', 'used to address both goals .', 'However , only is it computationally expensive , but there', 'are significant issues associated with ligand ranking [5] , [7] .', 'Thus , fast and accurate', 'methods for both binding pose prediction and ligand ranking need to be developed .', 'With the rapid increase in the number of experimentally solved protein structures ,', 'protein homology modeling has become a powerful tool in modern structural biology', '[8] , [9] .', 'Comparative modeling methods identify homologous', 'protein structures and use them as structural templates to model the target protein', 'of unknown tertiary structure .', 'Using a high sequence identity template with a clear', 'evolutionary relationship to the target , the modeled target structure can have a', 'root-mean-square-deviation , RMSD , from the native structure <2 Å', '[10] .', 'In', 'the “twilight zone” of sequence identity [11] , structural information', 'extracted from weakly homologous structure templates identified by threading is', 'sufficient to provide approximately correct 3D models for a significant fraction of', 'protein targets [12] , [13] .', 'In contrast to protein structure prediction ,', 'information from related 3D structures is rarely used in the large-scale modeling of', 'protein-ligand complexes .', 'One example of an approach that employs such information is CORES , an automated', 'method for building three-dimensional protein-ligand complexes [14] .', 'CORES directly utilizes', 'the conformation and binding pose of key structural elements of the target ligand ,', 'termed “molecular frameworks” , found in templates that are', 'closely related to the protein target .', 'Its practical utility was demonstrated on a', 'set of protein kinases in which ligands containing related frameworks were found to', 'bind in the same orientation .', 'A similar approach designed specifically for kinases ,', 'kinDOCK , performs ligand comparative docking by using a kinase family profile to', "align the related kinase-ligand complexes onto the target kinase's", 'structure and then directly transfers the ligand coordinates [15] .', 'KinDOCK typically', 'docks target ligands into the kinase binding pocket within a 2 Å RMSD from', 'the crystal structure .', 'Moreover , an original clustering procedure based on the', 'binding pose similarity was proposed to highlight the structural similarities and', 'differences within a set of multiple X-ray structures complexed with different', 'ligands [16] .', 'Other examples of ligand docking studies that', 'utilize structural information extracted from closely related protein-ligand', 'complexes include the analysis of cathepsin inhibitor specificity [17] , the', 'examination of carbohydrate recognition by the viral VP1 protein [18] , screening for selective bacterial sirutin inhibitors', '[19]', 'and the design of small molecule inhibitors of the macrophage migration inhibitory', 'factor [20] .', 'Typically , modeling templates for the target ligands are extracted from 3D', 'structures of small molecules complexed to closely related proteins .', 'In our previous study , we observed that evolutionarily remotely related proteins', 'identified by threading often share a common ligand-binding site [21] .', 'Both the localization of the binding site and chemical properties of bound ligands', 'are strongly conserved .', 'This forms the basis of the FINDSITE binding site', 'prediction/protein functional inference/ligand screening algorithm [21] .', 'Furthermore , we found that a pocket-specific potential of mean force derived from', 'known protein-ligand complexes identified for a given target sequence by threading', 'is often more specific than generic knowledge-based potentials derived from', 'ligand-protein complexes found in the PDB [22] .', 'This enhanced', 'specificity suggests that the binding mode and protein-ligand interactions in', 'distantly related protein families are conserved during evolution .', 'To confirm this', 'hypothesis , here , we present the results of ligand binding mode analysis of', 'evolutionarily distant proteins identified by state-of-the-art threading methods', '[23] .', 'The ligands that bind to the common binding site', 'contain a set of strongly conserved anchor functional groups as well as a variable', 'region that imparts specificity to a particular family member .', 'Furthermore , the', 'degree of sequence and structure conservation of residues in contact with the ligand', 'anchor functional groups are higher than those contacting ligand variable regions .', 'Exploiting these observations , we develop FINDSITELHM ( LHM stands for', 'Ligand Homology Modeling )', 'that employs structural information extracted from weakly related proteins to', 'perform rapid ligand docking and ranking by homology modeling; we compare its', 'accuracy to classical ligand docking/ranking approaches [4] , [22] , [24] .'], ['The protocol followed in this study is a direct extension of FINDSITE [21] ,', 'a threading-based method for ligand-binding site prediction and functional', 'annotation that detects the conservation of functional sites and their', 'properties in evolutionarily related proteins .', 'For a given target sequence ,', 'FINDSITE identifies ligand-bound template structures from a set of distantly', 'homologous proteins ( here , we limit ourselves to target proteins having', '<35% sequence identity to their closest template , but this', 'arbitrary restriction would be removed in real world predictions ) recognized by', 'the PROSPECTOR_3 threading approach [23] and superimposes', "them onto the target's ( experimental or predicted ) structure using the", 'TM-align structure alignment algorithm [25] .', 'Binding pockets are', 'identified by the spatial clustering of the center of mass of template-bound', 'ligands that are subsequently ranked by the number of binding ligands .', 'For each target protein , the template-bound ligands that occupy a top-ranked ,', 'predicted binding site are clustered using the SIMCOMP chemical similarity score', '[26] .', 'The “anchor” substructure is', 'then identified in each cluster as described in Methods .', 'First , we examine the', 'anchor substructure size relative to the average molecule size .', 'Applying the', 'approach to a representative benchmark set of 711 ligand-protein complexes', '( where the target proteins have pairwise sequence identity to their templates', '<35% , see Methods ) , as shown in Figure 1 , in most cases , at least', '50% of a ligand is comprised of an anchor region whose functional', 'groups are conserved in >90% of the template ligands .', 'Those', 'clusters in which the anchor region is smaller than 50% of the ligand', 'are mostly short oligosaccharides , with a sugar monomer identified as a common', 'substructure .', 'This also explains the high standard deviation in the average', 'ligand molecule size .', 'For some difficult cases , our graph isomorphism analysis', "didn't provide a sufficient number of atomic equivalences to recognize", 'a common substructure .', 'In contrast , those targets near the diagonal have an', 'anchor equivalent to the average molecule size and represent strongly conserved', 'ligands with little chemical variability; e . g . hemes .', 'In addition , there are', 'targets with a very small number of templates , all having very similar ligands .', 'Nonetheless , for the majority of targets , a well-defined anchor substructure', 'with a co-occurring variable region is detected .', 'Having identified the anchor substructure , we next investigate the structural', 'conservation of its binding mode .', 'Figure 1 ( inset plot ) shows the histogram of the average pairwise', 'RMSD among the anchor groups upon global superposition of the template proteins .', 'Note that the properties of the native ligand are not used in any way to', "identify the anchor region's properties .", 'Clearly , in most cases , the', 'average pairwise RMSD is <2 . 5 Å .', "We next examine the properties of the protein's ligand-binding region .", 'Given the chemical conservation of the anchor substructure as well as the strong', "structural conservation of it's binding mode , for binding residues , one", 'would expect that residues contacting ligand anchor groups are more conserved', 'than average .', 'The degree of sequence and structure conservation was calculated', 'for consensus binding residues ( CBRs ) , defined as residues contacting a ligand', 'in at least 25% of the threading templates .', 'This criterion was', 'previously found to maximize the overlap between predicted and observed binding', 'residues [21] and provides sufficient statistics to', 'calculate the sequence and structural features of binding residues .', 'We used a', 'probability threshold to define anchor/non-anchor CBRs based on the', 'protein-ligand contacts extracted from the threading templates .', 'The probability', 'of a residue to be an anchor residue simply corresponds to the fraction of', 'contacts formed by all residues in the equivalent position in the template', 'structures with anchor functional groups of bound ligands .', 'Differences in the', 'degree of sequence and structure conservation between anchor and non-anchor CBRs', 'were calculated on increasing the probability threshold from 0 . 1 to 0 . 9 using', "Student's t-test for independent samples .", "Shannon's", 'information entropy is used to measure the sequence variability at a particular', 'position in a target protein ( see Methods ) .', 'Analysis of the sequence entropy', 'revealed a significantly higher sequence conservation of residues in contact', 'with the anchor functional groups than those in contact with ligand variable', 'regions ( Figure 2A ) .', 'Next ,', 'we analyzed the structural features of CBRs in terms of the experimental', 'B-factors that reflect local mobility [27] and find that', 'the B-factors of residues in contact with the anchor region of the ligands are', 'significantly lower ( Figure 2B and', '2C which shows the B-factors of the Cαs and side chain heavy', 'atoms , respectively ) .', 'The conservation of the anchor-binding pose is consistent', 'with the relatively lower B-factors observed for the residues in spatial', 'proximity to the anchor functional groups .', 'These results differ significantly', 'from random ( Figure 2D ) .', 'Given that Figure 1 ( inset )', 'strongly suggests that the localization of the anchor substructure and its', 'internal conformation is conserved , we developed FINDSITELHM , a very', 'simple , rapid approach for ligand binding pose prediction .', 'Using the', 'consensus-binding mode of the anchor substructure , we align the ligand of', 'interest to the anchor region and then , optionally , minimize the ligand', 'conformation to remove steric clashes .', 'This procedure can be thought of as', '“ligand docking by homology modeling” .', 'Here , only weakly', 'related template proteins ( <35% sequence identity to the', 'target ) selected by threading were used to derive the consensus anchor-binding', 'mode .', 'In Table 1 , using', 'the crystal structures as the target receptors for ligand docking for the 711', 'ligand-protein set , the results are compared to three established ligand docking', 'approaches [4] , [22] , [24] in', 'terms of the heavy atom RMSD from the crystal structure .', 'Target proteins are', 'divided into three subsets with respect to the coverage of the predicted anchor', 'substructure .', 'For the first subset ( full coverage ) that consists of proteins for', 'which a portion of their target ligands cover at least 90% of the', 'functional groups in the predicted anchor substructure , simple ligand', 'superposition is quite successful and outperforms regular ligand docking', 'approaches .', 'For these cases , using all-atom minimization with Amber [28] ,', 'the predicted binding mode can be refined to an average RMSD from the crystal', 'structure of ∼2 . 5 Å .', 'An example of successful refinement is', 'presented for the human fibroblast collagenase in Figure 3 , where the final ligand heavy atom', 'RMSD is 0 . 63 Å .', 'In contrast , the RMSD from AutoDock is 2 . 77', 'Å .', 'The second subset ( partial coverage ) comprises target ligands that do not fully', 'cover any of the predicted anchor substructure .', 'Here , the average RMSD of the', 'binding mode predicted by FINDSITELHM is higher than AutoDock and is', 'comparable to Q-Dock and LIGIN .', 'However , it is still better than random ligand', 'placement .', 'Finally , if none of the predicted anchor substructures are even', 'partially covered by a target ligand ( low coverage ) , the results of docking', 'using FINDSITELHM are indistinguishable from random .', 'Here ,', 'traditional ligand docking approaches , particularly AutoDock , give much better', 'results .', 'In addition to anchor structure coverage , the performance of', 'FINDSITELHM depends on the overall accuracy of binding pocket', 'prediction and the conservation of the anchor-binding mode; this is discussed in', 'further detail below and presented in Figure 4 , see below .', 'Here , we note that using', 'the fraction of the anchor region that is aligned for a given ligand ( Figure 4A ) , or the average', 'pairwise RMSD of the anchor ligand functional groups ( Figure 4B ) , we can predict the expected', 'accuracy of binding pose prediction without knowing the experimental result .', 'Not', 'surprisingly ( Figure 4C ) ,', 'when the accuracy of the binding pocket prediction as provided by FINDSITE', 'improves , the accuracy of the ligand pose prediction by FINDSITELHM', 'also improves .', 'Weakly homologous protein models frequently have significant structural', 'inaccuracies in side-chain and backbone coordinates and thus , are much more', 'challenging targets for ligand binding pose prediction .', 'The performance of', 'FINDSITELHM , AutoDock , Q-Dock and LIGIN in ligand docking when', 'protein models are used as the target receptors was assessed for the Dolores', 'dataset of 205 proteins [22] , [29]; the average', 'Cα RMSD to native of these protein models is 3 . 7 Å .', 'Table 2 presents ligand', 'docking results using crystal structures as well as weakly homologous protein', 'models in terms of the fraction of recovered binding residues and specific', 'native contacts .', 'Considering the complete dataset and receptor crystal', 'structures , the accuracy of FINDSITELHM is slightly lower than', 'AutoDock and Q-Dock .', 'This is because the predicted anchor substructure was fully', 'covered ( ≥90% ) by the target ligand only for 62 . 4%', 'of the receptors; partial ( ≥50% and <90% )', 'and low ( <50% ) coverage of the anchor substructure was found', 'for 25 . 4% and 12 . 2% of the targets , respectively .', 'This', 'partly reflects the fact that the placement of the ligand variable region has a', 'random component that diminishes the overall accuracy .', 'Consistent with the', 'decrease in ligand RMSD on minimization , the fraction of binding residues and', 'native contacts increases .', 'In contrast , for protein models , FINDSITELHM recovered more binding', 'residues and specific native contacts than both all-atom docking approaches ,', 'AutoDock and LIGIN .', 'Considering only the most confident cases for which FINDSITE', 'was likely to predict the binding pocket center with ≤4 Å', 'accuracy ( “Easy” targets ) and the predicted anchor', 'substructure fully ( partially ) covered by the target ligand , the fraction of', 'binding residues and specific native contacts recovered by', 'FINDSITELHM is 0 . 66 ( 0 . 61 ) and 0 . 49 ( 0 . 43 ) , respectively .', 'However ,', 'now the all-atom minimization procedure applied to the binding poses predicted', 'by FINDSITELHM caused a loss of the specific native contacts .', 'This', 'reflects the fact that structure adjustments are required to remove the', 'repulsive ligand-residue interactions that are not accommodated by simple', 'minimization .', 'Nevertheless , these results represent a significant improvement', 'over traditional all-atom docking against modeled receptor structures .', 'We also', 'note the high sensitivity of all-atom docking approaches to the quality of the', 'receptor structures; for weakly homologous protein models , the performance of', 'AutoDock and LIGIN is no better than random ligand placement into the predicted', 'binding sites .', 'The performance of Q-Dock for protein models was notably higher ,', 'since it was explicitly designed to deal with structural inaccuracies in', 'predicted receptor models .', 'Finally , in contrast to classical ligand docking', 'approaches , FINDSITELHM is computationally less expensive , and', 'typically requires less than a minute of CPU time ( see Table', 'S1 ) .', 'An interesting question that is very important from the practical point of view ,', 'is when should we expect a successful binding mode prediction by using ligand', 'docking by homology modeling ?', 'In addition to the coverage of an anchor structure', 'that clearly impacts docking accuracy ( Figure 4A ) , we also investigated the', 'relationship between pocket prediction accuracy , expressed as the distance', 'between the predicted pocket center and the geometric center of the native', 'ligand , the conservation of anchor binding mode in terms of the average pairwise', 'RMSD of the anchor functional groups , and the accuracy of FINDSITELHM', 'binding mode prediction assessed by the heavy atom RMSD from the crystal ligand', 'pose .', 'As expected , the average accuracy of the binding mode prediction by', 'FINDSITELHM decreases with decrease in the degree of the', 'conservation of the anchor substructure ( Figure 4B ) .', 'The RMSD of the predicted', 'ligand-binding pose is <2 Å on average for highly conserved', 'anchor substructures whose pairwise RMSD is <2 Å .', 'For', 'moderately conserved anchor substructures with a pairwise RMSD of 2–4', 'Å , the RMSD of the predicted ligand-binding mode is <3', 'Å in most cases .', 'Finally , accompanied by weak ( >4 Å )', 'structural conservation of an anchor , docking accuracy drops to >3', 'Å on average .', 'In addition , the drop off in ligand binding pose', 'prediction correlates with the overall accuracy of binding pocket prediction by', 'FINDSITE ( Figure 4C ) .', 'The', 'most accurate ligand binding poses were obtained for precisely detected pockets ,', 'where the pocket center was predicted within 2 Å from the geometric', 'center of the native ligand .', 'Considering the structural conservation of the', 'derived anchor substructure , its coverage by a target ligand and the FINDSITE', 'confidence index for pocket detection [21] , ( all properties', 'which can be calculated without knowledge of the native binding', 'pose ) , one can roughly estimate the quality of the performance of', 'FINDSITELHM in ligand binding pose prediction .', 'The results of the application of FINDSITELHM to glutathione', 'S-transferase ( PDB-ID: 1a0f ) , MTA phosphorylase ( PDB-ID: 1sd2 ) and lysine', 'aminotransferase ( PDB-ID: 2cjd ) are presented .', 'Figures 5–10 present the common ligand anchor substructures/variable groups', 'identified from weakly homologous threading templates for these 3 proteins .', 'In', 'Figure 11 , the degree of', 'sequence and structure conservation of amino acid residues for these proteins is', 'projected onto the target protein surface and compared to a random distribution .', 'In Figures', '12–14 , using the target crystal', 'structure , the results of flexible ligand docking by FINDSITELHM', '( including refinement ) are compared to ligand binding poses predicted by', 'classical docking approaches and the consistently better performance of', 'FINDSITELHM is demonstrated .', 'In the case of Figure 14 where the RMSDs of LIGIN and random', 'pose prediction are the same as FINDSITELHM , the', 'pyridoxal-5′-phosphate moiety is clearly better placed by', 'FINDSITELHM .', 'All have the same RMSD mainly due to the incorrect', 'placement of the variable region .', 'Recently , a detailed picture of the evolution and diversification of enzyme', 'function was drawn from the analysis of conservation of substrate substructures', 'in 42 major enzyme superfamilies [30] .', 'Based on graph', 'isomorphism analysis , highly conserved substructures were identified in all', 'substrates of a particular enzyme superfamily .', 'For the remaining substrate', 'substructures , called reacting substructures , substantial variation in chemical', 'properties within the superfamily was found .', 'Systematic analysis of the', 'substrates in 42 major SCOP [31] enzyme superfamilies revealed chemically', 'conserved patterns that typify individual superfamilies [30] .', 'This approach is', 'very similar in spirit to FINDSITELHM; both demonstrate how', 'evolutionary pressure directs the evolution of protein molecular function .', 'The', 'structural and chemical patterns of enzyme substrates , or small ligands in', 'general , have been conserved during evolution due to the strong conservation of', 'the structural and chemical features of the binding site residues .', 'We next analyzed the overlap between the conserved substrate substructures ( CSSs )', 'identified at the SCOP superfamily level [30] and the anchor', 'regions in ligands bound to evolutionarily related proteins selected by', 'threading .', 'The results presented in Figure 15 show that the highly conserved substructures of the enzyme', 'substrates identified by Babbitt and colleagues [30] to a large extent', 'overlap with the anchor substructures detected by our threading-based approach;', 'in over 70% of the cases , the anchor substructure covers at least', "70% of CSS's atoms .", 'Detailed results obtained for', '4-α-glucanotransferase from T . litoralis ( PDB-ID: 1k1w )', 'and D-xylose isomerase from Arthrobacter sp .', '( PDB-ID: 1die ) are', 'presented in Tables S2 and S3 , respectively .', 'We find that the highly', 'conserved substructures of the enzyme substrates frequently overlap with the', 'conserved anchor substructures detected by our threading-based approach .', 'The set', 'of ligands that bind to the common binding site in distantly evolutionarily', 'related proteins contain a set of strongly conserved', '“anchor” functional groups and', '“variable” regions that account for a specificity toward a', 'particular family member .', 'As a consequence of the ligand clustering procedure that precedes anchor', 'identification , the anchor substructures typically contain more atoms than CSSs', 'and are not confined to enzymes .', 'Both features are important for practical', 'application in ligand docking by homology modeling , as demonstrated by', 'FINDSITELHM simulations , where the consensus anchor-binding mode', 'is used as a reference framework for the superposition of a query ligand .', 'Furthermore , common anchor substructures are observed across ligands bound to', 'weakly related proteins that belong to more than one superfamily .', 'These subtle', 'evolutionary relationships detected by sensitive threading techniques [32] , [33] are of paramount', 'importance for novel biopharmaceutical discovery that could be accounted for to', 'identify potential off-site drug targets and reduce side effects .', 'HIV-1 protease plays a crucial role in the life cycle of HIV [34] , [35]; thus , it is an', 'important drug target for AIDS treatment with a number HIV-1 protease inhibitors', 'identified [36] , [37] .', 'Several ( Table 3 ) are FDA-approved', 'anti-HIV drugs .', 'Here , we selected HIV-1 protease as an example to demonstrate', 'the performance of FINDSITELHM in ligand-based virtual screening', 'using the coverage of anchor substructures as a simple scoring function .', 'The performance of FINDSITELHM alone and in combination with FINDSITE', 'in virtual screening for HIV-1 protease inhibitors is presented in Figure 16 .', 'Both FINDSITE and', 'FINDSITELHM perform considerably better than a random ligand', 'selection .', 'The molecular fingerprints constructed by FINDSITE recovered slightly', 'more known active compounds in the top-ranked fraction of the screening library', 'than anchor-based FINDSITELHM; the enrichment factor calculated for', 'the top 1% ( 10% ) is 27 . 0 ( 6 . 8 ) and 23 . 3 ( 5 . 9 ) for FINDSITE', 'and FINDSITELHM , respectively .', 'Clearly , fusion by ranks outperforms', 'the individual scoring functions with the enrichment factor of 38 . 1 ( 7 . 3 ) for', 'the top 1% ( 10% ) of ranked ligands .', 'These results suggest', 'that the anchor-based approach is able to detect active compounds for which the', 'fingerprint-based method assigns relatively low score .', 'Furthermore , using the', 'combined FINDSITE/FINDSITELHM approach , 4 ( 7 ) out of 10 FDA-approved', 'HIV-1 protease inhibitors are found in the top 1% ( 5% ) of', 'the screening library ( Table', '3 ) .'], ['Conservation of protein sequence and structural patterns is widely used to study', 'protein molecular function [38]–[40] .', 'Indeed , the structural', 'and chemical characteristics of a binding site are important for understanding', 'ligand selectivity and cross-reactivity [41] , [42] .', 'In that regard , our', 'sequence entropy analysis suggests that residues contacting anchor functional groups', 'have been subjected to higher evolutionary conservation pressure than those', 'contacting ligand variable regions .', 'Furthermore , the conservation of the', 'anchor-binding pose is consistent with the relatively low experimental B-factors', 'observed for residues contacting anchor functional groups .', 'The significantly higher', 'structural plasticity of variable region binding residues could reflect the', 'different types/sizes of functional groups found in the ligand variable', 'substructures that might be responsible for ligand specificity for particular', 'protein family members .', 'Binding site analysis also has practical implications .', 'In the simplest case , using', 'the ligand binding modes extracted from closely related structures and incorporated', 'as spatial restraints in protein structure modeling provides better homology models', 'of protein binding sites [43] .', 'In large-scale computational experiments', 'involving ligand docking , using the AnnoLyze approach the transfer of ligands from', 'known structures of closely related protein-ligand complexes is an attractive', 'alternative to CPU-expensive , classical ligand docking approaches [44] .', 'Here , we have shown that this idea is in fact more general and applies to', 'evolutionarily distant proteins .', 'Indeed , evolution provides a type of signal', 'averaging to identify the essential features associated with ligand binding .', 'This', 'insight can be profitably exploited in a variety of contexts .', 'For example , for evolutionary distant proteins , we identify the subset of ligands', 'whose pose is conserved , viz . the anchor region .', 'Then , based on the observation that', 'across a set of weakly related proteins , not only is the chemical identity of anchor', 'functional groups strongly conserved but also the anchor binding mode , with an', 'average pairwise RMSD <2 . 5 Å in most cases .', 'FINDSITELHM', 'uses the consensus binding mode of an anchor substructure as the reference', 'coordinates to perform rapid flexible ligand docking by superposition .', 'This results', 'in an average ligand heavy atom RMSD from native of 2 . 5 Å for those', 'ligands that contain a significant portion of the anchor region .', 'Moreover , for', 'predicted protein structures , with considerably less CPU time ,', 'FINDSITELHM outperforms all-atom ligand docking approaches in terms of', 'the fraction of recovered binding residues and specific native contacts .', 'The accuracy of FINDSITELHM is affected by several factors: First , for a', 'given target , the set of evolutionarily related template structures needs to be', 'identified .', 'Given the improvements in threading approaches [23] , [45] and the completeness of the', 'fold library [46] , one can expect to obtain a set of templates for the', 'majority of single domain targets .', 'Next , the docking performance of', 'FINDSITELHM is well correlated with the overall accuracy of binding', 'pocket prediction by FINDSITE .', 'Typically , high accuracy in ligand binding pose', 'prediction requires the binding site to be precisely detected within a distance of 2', 'Å .', 'This level of accuracy in pocket prediction is usually achieved for', 'Easy targets , as classified by FINDSITE [21] .', 'Finally , the', 'average accuracy of the binding mode prediction by FINDSITELHM decreases', 'with the decrease in the coverage of the anchor substructure by the target ligand as', 'well as with the decrease in the degree of the anchor structural conservation .', 'Here ,', 'the growing number of protein crystal structures solved in the complexed state with', 'chemically diverse small organic molecules expands the pool of suitable targets for', 'FINDSITELHM .', 'It is noteworthy from the practical point of view that', 'all these properties can be calculated during the modeling procedure , without', 'knowing the native binding pose .', 'Thus the expected accuracy of', 'FINDSITELHM in ligand binding pose prediction can be estimated with', 'fairly high confidence .', "Also as shown for HIV-1 protease , using just the target protein's sequence", 'as input , FINDSITE/FINDSITELHM can efficiently and rather accurately rank', 'a large ligand library .', 'Since for the majority of gene products , at least weakly', 'homologous proteins can be identified in structural databases by current threading', 'methods [23] and approximately correct protein models can be', 'generated by protein structure prediction techniques [10] , [12] , [13] , FINDSITELHM', 'offers the possibility of proteome-scale structure-based virtual screening for novel', 'biopharmaceutical discovery .', 'This would have a great advantage over just screening', 'single proteins .', 'It affords the possibility of identifying lead compounds with', 'desired selectivity that could be further exploited at the outset of the drug', 'development process to reduce side effects .', 'We note that similarity in global fold alone is usually insufficient for effective', 'function interference and results in a high false positive rate [47] .', 'For', 'that reason , the most effective function prediction methods , such as ProFunc [48] ,', 'AnnoLite [44] or Mark-Us [49] typically combine', 'structure- and sequence-based techniques .', 'In that respect , an important component of', 'FINDSITE/FINDSITELHM is the template selection by threading that', 'employs a strong sequence profile term [23] .', 'This allows the', 'detection of evolutionarily distant homologues [21] with clear', 'functional relationships to the protein of interest not only in terms of the', 'localization of the binding site , but also in the detailed chemical and structural', 'aspects of ligand binding , particularly those that impart binding specificity .', 'Thus ,', 'threading provides a richness to functional annotation that to date was not fully', 'exploited .'], ['High quality protein–ligand complex X-ray structures were taken from', 'the Astex diverse set used to validate the GOLD docking algorithm [50]', 'and from a non-redundant Q-Dock dataset [22] .', 'In the Astex', 'set , we excluded complexes in which the binding site is formed by more than one', 'protein chain .', 'From the Q-Dock set , we exclude proteins with', '>35% sequence identity to any protein in the Astex set .', 'We', 'only include proteins for which at least 5 ligand-bound weakly homologous', 'threading templates can be identified by protein threading and the binding', 'pocket can be predicted by FINDSITE [21] within 4 . 5', 'Å from the bound ligand; this represents about 67% of', 'protein targets .', 'The final dataset consisting of 711 complexes is found at', 'http://cssb . biology . gatech . edu/skolnick/files/FINDSITELHM .', 'In addition to the crystal structures used as the ta
['Introduction', 'Results', 'Discussion', 'Methods']
['Ligand virtual screening is a widely used tool to assist in new pharmaceutical', 'discovery .', 'In practice , virtual screening approaches have a number of', 'limitations , and the development of new methodologies is required .', 'Previously ,', 'we showed that remotely related proteins identified by threading often share a', 'common binding site occupied by chemically similar ligands .', 'Here , we demonstrate', 'that across an evolutionarily related , but distant family of proteins , the', 'ligands that bind to the common binding site contain a set of strongly conserved', 'anchor functional groups as well as a variable region that accounts for their', 'binding specificity .', 'Furthermore , the sequence and structure conservation of', 'residues contacting the anchor functional groups is significantly higher than', 'those contacting ligand variable regions .', 'Exploiting these insights , we', 'developed FINDSITELHM that employs structural information extracted', 'from weakly related proteins to perform rapid ligand docking by homology', 'modeling .', 'In large scale benchmarking , using the predicted anchor-binding mode', 'and the crystal structure of the receptor , FINDSITELHM outperforms', 'classical docking approaches with an average ligand RMSD from native of', '∼2 . 5 Å .', 'For weakly homologous receptor protein models , using', 'FINDSITELHM , the fraction of recovered binding residues and', 'specific contacts is 0 . 66 ( 0 . 55 ) and 0 . 49 ( 0 . 38 ) for highly confident ( all )', 'targets , respectively .', 'Finally , in virtual screening for HIV-1 protease', 'inhibitors , using similarity to the ligand anchor region yields significantly', 'improved enrichment factors .', 'Thus , the rather accurate , computationally', 'inexpensive FINDSITELHM algorithm should be a useful approach to', 'assist in the discovery of novel biopharmaceuticals .']
['As an integral part of drug development , high-throughput virtual screening is a', 'widely used tool that could in principle significantly reduce the cost and time', 'to discovery of new pharmaceuticals .', 'In practice , virtual screening algorithms', 'suffer from a number of limitations .', 'The high sensitivity of all-atom ligand', 'docking approaches to the quality of the target receptor structure restricts the', 'selection of drug targets to those for which high-quality X-ray structures are', 'available .', 'Furthermore , the predicted binding affinity is typically strongly', 'correlated with the molecular weight of the ligand , independent of whether or', 'not it really binds .', 'To address these significant problems , we developed', 'FINDSITELHM , a novel threading-based approach that employs', 'structural information extracted from weakly related proteins to perform rapid', 'ligand docking and ranking that is very much in the spirit of homology modeling', 'of protein structures .', 'Particularly for low-quality modeled receptor structures ,', 'FINDSITELHM outperforms classical all-atom ligand docking', 'approaches in terms of the accuracy of ligand binding pose prediction and', 'requires considerably less CPU time .', 'As an attractive alternative to classical', 'molecular docking , FINDSITELHM offers the possibility of rapid', 'structure-based virtual screening at the proteome level to improve and speed up', 'the discovery of new biopharmaceuticals .']
['biophysics/biomacromolecule-ligand interactions', 'biochemistry/biomacromolecule-ligand interactions', 'biochemistry/drug discovery']