Creating a pharmacophore from a single protein-ligand complex

Transcription

1 CARD 1 Creating a pharmacphre frm a single prtein-ligand cmplex basic minutes User Cntrls Advanced cntrls (pt.) Macrmlecule view Active site view Dwnlad PDB file using -letter cde Discver crrect ligand Fcus n active site and switch t active site view Check and eventually crrect ligand chemistry Create a structure-based pharmacphre Ligand bx Create pharmacphre (buttn r menu) OR: Create pharmacphre fr Catalyst/MOE/ Phase Change bnd type Change atm type Create new bnd Mve t cre Mve t envirnment Cmpund switching Descriptin: Type 1ke6 in upper right area f screen and press buttn Dwnlad 1ke6 [1]. The prtein will be dwnladed and displayed (this is macrmlecule view ) [2]. Click n yellw bx within prtein representing ligand - an animated zm begins ending in [3]. Since ligand structures in PDB nly cntain incmplete infrmatin, yu shuld always check wher all bnds are typed crrectly. Bnd types can be changed by selecting a bnd (by clicking n it eir in 2D r 3D view) and by using retype bnd buttn [] r keys 1, 2, r 3. Once ligand is chemically crrect, create a pharmacphre by pressing Ctrl-F9 (Cmd-F9 n OS X) []. Where t g frm here: Pharmacphre mdeling: Creating shared and merged feature pharmacphres Custmizing pharmacphre creatin preferences Virtual screening 1 2 3

2 CARD 2 Pharmacphre mdeling: Create shared & merged feature pharmacphres using chemical features frm three PDB structures medium 1 minutes User Cntrls Advanced cntrls (pt.) Macrmlecule view Active site view Alignment view Dwnlad PDB File 1ke6 Check & crrect ligand Create pharmacphre Add ligand and pharmacphre t alignment view Repeat last steps fr 1ke7 and 1ke8 Create a shared feature pharmacphre frm 3 pharmacphres Align three ligands t new pharmacphre Ligand bx Create pharmacphre (buttn r menu) Add mlecule t alignment Add pharmacphre t alignment Generate shared feature pharmacphre (by features) Generate merged feature pharmacphre Set reference element Center all structures Alignment cntext menu Descriptin: Type 1ke6 in upper right area f screen and press buttn Dwnlad 1ke6. The prtein will be dwnladed and displayed in macrmlecule view. Click n yellw bx t fcus n ligand LS2201. Check ligand fr its crrectness. Press ctrl-f9 t create a pharmacphre (cmd-f9 n OS X). Use data exchange widget and select add mlecule t alignment view [1a] and mlecule will be added t alignment view, increasing a cunter in crrespnding tab. Use data exchange widget and select add pharmacphre t alignment view [1b] t d same with pharmacphre. Repeat se steps with 1ke7, and 1ke8. Finally, yu shuld have six elements in alignment view. Click n Alignment tab t wrk with m. Select three pharmacphres in alignment list using Ctrl-click r multiple selectin [2], and press Generate shared feature pharmacphre (by features) buttn [3]. A new pharmacphre called Shared [LS2201-1, LS3201-1, LS2-1] is appended t list. Mark this element and make it reference element using Set reference buttn, which will change display style f this element t red and bld. Nw use ctrl key (cmd n OS X) t additinally select mlecules identified by blue icns []. Additinally, hide three pharmacphres derived frm 1ke6, 1ke7, and 1ke8 by clicking n eye symbls n right next t se alignment entries []. Nw press align buttn. Yu will see three ligands aligned t shared feature pharmacphre derived frm three PDB cmplexes []. Where t g frm here: Custmizing pharmacphre creatin preferences Virtual screening in LigandScut 1 2 3

3 CARD 3 Virtual screening and cmparing active/inactive hit retrieval rates advanced 1 minutes User Cntrls Advanced cntrls (pt.) Macrmlecule view Active site view Create pharmacphre fr virtual screening Initiate virtual screening Refine pharmacphre mdel Repeat virtual screening Ligand bx Create pharmacphre (buttn r menu) Initiate virtual screening Virtual screening with mitted features Descriptin: Virtual screening aims at maximum enrichment f active cmpunds in a hit list. Therefre, such methds are usually validated by assessing accuracy f discriminatin between actives and decys. A gd pharmacphre mdel will be able t identify a significant prtin f knwn active cmpunds and as few decys as pssible. In this tutrial, tw different types f precalculated multicnfrmatinal databases are available: A library cntaining a selectin f knwn active cmpunds and a library f decys. Type 1ke7 in upper right area f screen, press buttn Dwnlad 1ke7. Create a pharmacphre f prteinligand cmplex as described in LigandScut tutrial sheet 1 Creating a pharmacphre frm a single prtein-ligand cmplex [1]. Perfrm virtual screening using a multi-cnfrmatinal database with knwn active cmpunds (filename cdk2- ligands.ldb ) fr screening using Screen pharmacphre against external library item lcated in Pharmacphre menu [2]. Virtual screening will be initiated after selecting library file (press butten with three dts t pen a file dialg) and number f mittable features [3]. Since initial pharmacphre was derived frm all interactins f nly ne single ligand, it is rar restrictive and retrieves nly few hits. Dwnsize number f features by deleting less imprtant features [] r increase tlerances f features by selecting a feature and n chsing Pharmacphre -> Increase Selected Feature Tlerance frm menu. Cmpare rati f active cmpunds and decys (library filename cdk2-decys.ldb ) retrieved by yur pharmacphre in rder t estimate its predictive pwer []. Furr adapt pharmacphre in rder t imprve rati f actives and decys btained by virtual screening: Remve nn-essential chemical features and exclusin vlume spheres in rder t increase sensitivity; adapt size f feature spheres by pressing ctrl-shift-plus r crrespnding menu entry ( Pharmacphre - Decrease Selected Feature Tlerance ) r use pharmacphre frm tutrial card 2 using data exchange widgets [6]. Where t g frm here: Pharmacphre mdeling: Creating shared and merged feature pharmacphres Custmizing pharmacphre creatin preferences Analyzing screening results

4 CARD Analyze LigandScut pharmacphre screening results medium 10 minutes Prerequisites: Virtual screening hit list User Cntrls Advanced cntrls (pt.) Library view (spreadsheet) Hierarchy view Create r imprt screening results Srt virtual screening hit list Inspect and cmpare hits Discver cntrls f library view Library view (Spreadsheet) Viewer cntrls Calculatin f Gaussin Shape Similarity Scre Descriptin: Perfrm virtual screening t btain a hit list as described n tutrial card Virtual screening and cmparing active/inactive hit retrieval rates r insert external screening results frm a library file via File - Insert menu entry. Results f virtual screening are depicted in library view [1]. Srt hit list by a single click n Pharm. Fit Scre table header [2]. Yu may invert srting rder by an additinal click. Select a cmpund f interest by single click n crrespnding table rw t be depicted in active site view tger with riginal ligand [3]. Multiple cmpunds are selected when hlding shift r ctrl (cmd n MAC OS X) while clicking. Keep cmpunds visible independent frm current selectin state by clicking eye symbl in table left t mlecule name in library view spreadsheet []. The visibility f cre mlecule (i.e., riginal ligand) and r items in active site view is tggled with eye symbl lcated in hierarchy view n right side next t mlecule name []. Select a custm clr fr cre mlecule using square icn next t eye icn in hierarchy view. Calculatin f Gaussian Shape Similarity Scre ffers furr functinality fr scring. Use Calculate Gaussin Shape Similarity Scre functin lcated in Library menu fr calculatin. A new prperty Shape Tanimt Scre will be added in library view and yu can use spreadsheet functinality fr srting and ranking as described abve. Where t g frm here: Pharmacphre mdeling: Creating shared and merged feature pharmacphres Custmizing pharmacphre creatin preferences 2 3 1

5 CARD View and rescre dcking pses with pharmacphres medium 10 minutes Prerequisites: dcking pses frm external surces User Cntrls Advanced cntrls (pt.) Library view (spreadsheet) Hierarchy view Dwnlad PDB File Switch t active site view Imprt dcking pses Analyze dcking pses Rescre dcking pses Align dcking pses t pharmacphre Discver viewer cntrls f library view Library view (Spreadsheet) Viewer cntrls Descriptin: Dwnlad 1ke7 and create pharmacphre f prtein-ligand cmplex. Imprt crrespnding dcking pses stred in MDL SDF file frmat (tutrial filename 1ke7-dcking.sdf ) via File - Insert menu entry [1]. Make sure that Align Mlecules Autmatically t Shwn Pharmacphre ptin lcated in Library menu is deactivated [2], which ensures that crdinates f dcking pses relative t prtein are preserved. The cmpunds are imprted t Ligand- Scut including all prperties given in SD file, such as dcking scre. Srt list f dcking pses by a single click n table header crrespnding t dcking scre [3]. Invert srting rder by an additinal click if necessary. LigandScut ffers fur different ptins fr rescring dcking pses based n 3D pharmacphre that is currently visible in LigandScut: The calculatin f pharmacphre fit f imprted dcking pses cnsidering r ignring excluded vlume spheres, and calculatin f pharmacphre fit f dcking pses after alignment t pharmacphre cnsidering r ignring excluded vlume spheres. Alignment t pharmacphre befre scring may be useful t ptimize dcking pses []. All fur mdes are accessed via Library menu. Yu can mdify pharmacphre by deleting features, adding excluded vlumes, etc. t reflect imprtant features f binding site that shuld be fulfilled by highest ranked hits. Alignment f imprted dcking pses t shwn pharmacphre is perfrmed autmatically upn selectin f a spreadsheet line (i.e. a single dcking pse) if Align Mlecules Autmatically t Shwn Pharmacphre ptin lcated in Library menu is activated. Pses may als be aligned t pharmacphre using LigandScut s alignment algrithm using Align buttn lcated in upper right crner f library view []. Where t g frm here: Custmizing pharmacphre creatin preferences 1 2 3

6 CARD 6 Espress: Create ligand-based 3D pharmacphres intermediate 1 minutes User Cntrls Advanced cntrls (pt.) Ligand-based Mdeling Imprt ligands Cluster ligand-set in 3D Generate ligand-based pharmacphre mdel 3D Clustering Generate ligand-based pharmacphre mdel (buttn r menu) Use different settings fr cnfrmatinal mdel generatin Use different settings fr clustering Use different settings fr ligand-based pharmacphre mdel generatin Descriptin: Open smiles file cntaining yur ligands ( cdk2.smi ) by chsing File-Open menu. The ligands are imprted and shwn in 2D n right side f screen [1]. Set all candidates fr mdel t training set by chsing Ligand-Set -> Flag Selected Mlecules as Training Set. Cluster ligand set accrding t ir 3D pharmacphre characteristics using cluster buttn n bttm f 3D view [2]. Keep default setting in fllwing dialg and start clustering prcess by pressing OK buttn. Espress will create a new clumn in ligand table called Cluster ID. Srt table using this clumn by clicking n clumn header Cluster ID. Set all mlecules with cluster ID 1 t test set by using table cntrls [3]. Nw click n buttn Create ligand-based pharmacphre [] using default values t start pharmacphre generatin. A ligand-based mdel will be created and displayed in 3D view. Feature patterns [] in table indicate, which features are met by which ligand. A click n a clred square indicates, which feature is linked t crrespnding square. The list n right lwer crner f screen shws several pharmacphre slutins: by selecting a different slutin, it is shwn in 3D view with ligands aligned accrdingly [6]. Yu can use data exchange cntrls n upper right crner f 3D view t mve currently shwn pharmacphre t r views (e.g. screening) r save it using File -> Save menu and selecting a file type that represents pharmacphres. Where t g frm here: Align structure-based pharmacphre mdels with ligand-based nes Virtual screening in LigandScut

7 CARD 7 Screen several databases using Screening Perspective intermediate minutes User Cntrls Advanced cntrls (pt.) Screening view Add pharmacphre mdel t screening view Lad virtual database Perfrm virtual screening Lad Screening Database (Buttn) Perfrm Screening (Buttn) nne Descriptin: In cntrast t virtual screening prcedure in Structure-based Mdeling perspective which is designed fr pharmacphre mdel refinement with respect t ligand-binding pcket prperties, Screening perspective prvides a fast and cnvenient way f perfrming multiple virtual screens with different screening databases against yur final pharmacphre mdel, which can be useful fr validatin. Imprt yur favrite pharmacphre mdel (e.g. ne created frm tutrial card 1 r 6) int Screening perspective eir by using Exchange Data Widget [1] r Open entry in File menu. Click n Lad Screening Database buttn [2] t lad screening database cdk2-ligands.ldb. Enable laded screening database fr screening by clicking n grey buttn left t screening database entry [3]. Nw, select pharmacphre mdel frm Pharmacphres list t make sure it is shwn in 3D view. Next t Lad Screening Database buttn, Perfrm Screening and Delete Pharmacphre buttn will be enabled. Click n Perfrm Screening buttn t initiate virtual screening. View results in table at bttm f screen []. Yu can add mre databases and pharmacphres fr furr virtual screening experiments. Please make sure that nly ne pharmacphre is visible befre yu start screening. Where t g frm here: Analyze screening results Advanced screening: ROC curves and mdel cmbinatin 1 2 3

8 CARD 8 Advanced screening: ROC curves and mdel cmbinatin advanced 1 minutes User Cntrls Advanced cntrls (pt.) Screening view Add pharmacphre mdels t screening view Lad virtual database Perfrm virtual screening Plt ROC curve Cmbine mdels by OR Lad Screening Database (Buttn) Perfrm Screening (Buttn) Plt ROC Curve (Buttn) Blean Expressin (Field) Descriptin: In cntrast t virtual screening prcedure in Structure-based Mdeling perspective which is designed fr pharmacphre mdel refinement with respect t ligand-binding pcket prperties, Screening perspective prvides means fr statistical evaluatin f screening perfrmance. Imprt structure-based pharmacphre mdels frm PDB entry 1ke7 (tutrial card 1) and ligand-based CDK2 pharmacphre (tutrial card 2) int Screening perspective eir by using Exchange Data Widget [1] r, alternatively, lad a saved versin using Open entry in File menu. Click n Lad Screening Database buttn [2] t lad screening database cdk2-ligands.ldb, and click same buttn again and lad database cdk2-decys.ldb. Click nce n tri-state selectr left t database names t mark cdk2-ligands.ldb as a database cntainaing actives (green database symbl), and twice n tri-state slectr left f cdk2-decys.ldb t mark this database as decys (red database symbl) [3]. Nw, select pharmacphre mdel 1ke7 frm Pharmacphres list t make sure it is shwn in 3D view. Click n Perfrm Screening buttn t initiate virtual screening []. After screening has finished, press Plt ROC Curve buttn [] t plt a ROC curve [6]. Nw select bth pharmacphres in Pharmacphres table. Numbers in curly brackets ( {1} and {2} ) indicate number f pharmacphre. Enter 1 r 2 int blean expressin field [7] indicating that a virtual hit in this screening run shuld eir fit int first (1) r t secnd (2) pharmacphre. Press screening buttn [] again t re-screen, and ROC curve buttn [] again t cmpare virtual screening statistics. View virtual hits in table at bttm f screen [8]. Yu can add mre databases and pharmacphres fr furr virtual screening experiments. Where t g frm here: Analyze screening results

9 CARD 9 12 LigandScut Mlecule Tables: Filtering & Exprt intermediate 1 minutes User Cntrls Advanced cntrls (pt.) IDBGEN Database generatr Structure-based view Table view Create a multicnfrmatinal database frm a smiles file Examine cmpunds Calculate physicchemical prperties Filter and exprt mlecular structures IDBGEN GUI Lad LDB file Calculate standard prperties Apply filter rules Exprt 2D structure files Exprt t Micrsft Excel Descriptin: Start IDBGEN GUI mlecular database generatr [1]. Chse a smiles file f mlecules as input file and specify utput file. Set icn FAST ptin fr cnfrmatin generatin [2]. After cmpletin, lad resulting ldb file int structure-based perspective and fcus n table view [3]. Calculate sme physicchemical parameters and mlecular descriptrs using Library -> Calculate Standard Prperties. Apply filters [, ] and view dynamically generated results. Where t g frm here: Exprt mlecular structures tger with calculated standard prperties t a 2D SDF file Exprt filtered cmpund list t a Micrsft Excel file

10 CARDCARD 10 a pharmacphre frm a single prtein-ligand cmplex In Creating Silic Fragment Screening with LigandScut advanced basic 1 minutes minutes Macrmlecular view Macrmlecule view Screening view Active site view Active site view User Cntrls User Cntrls Advanced cntrls (pt.) Advanced cntrls (pt.) Create structure based Ligand bx Re-dck resulting mlecules Dwnlad PDB file using Ligand bx Change bnd type pharmacphre mdel Create pharmacphre and minimize -letter cde Create pharmacphre Change atm type Switch t screening view Set features Re-scre new mlecules Discver crrect (buttnptinal r menu) Create bnd by Perfrm fragment screening Initiate virtual screening estimating free ligand OR: Create pharmacph Mve t cre energy Fcus n active site and Library review fr Catalyst/MOE/ binding Mve t envirnment Inject hits back int prtein f switch Cmpund switching binding site t active site view InjectPhase hits int prtein Exprt results Check and eventually cr Jin fragments t create binding site rect ligand chemistry new mlecules Create a single-click phar- Minimize hits Merge mlecules macphre Descriptin: Descriptin: Type 1ke6 in upper right area f screen, an press buttn Dwnlad 1ke6 [1]. The prtein will be dwnladed and displayed (this isright macrmlecule n Dwnlad yellw1ke7 bx [1], within prtein representing ligand - an Type 1ke7 in upper area f screen,view ) press [2]. Click buttn click n yellw bx, and generate a animated zm begins ending in [3]. Since ligand structures in PDB nly cntain incmplete infrmatin, yu shuld pharmacphre mdel. Mve mdel t Screening view. Lad tutrial fragment library (fragmentstutrial.ldb). Set left always check wher all bnds are typed crrectly. Bnd types can be changed by selecting a bnd (by clicking n it eir in three features as ptinal by selecting m (press Shift+left muse buttn and draw a rectangle arund m) and use 2D r 3D view) and by using retype bnd buttn [] r keys 1, 2, r 3. Once ligand is chemically crrect, create Pharmacphre -> Mark asctrl-f9 Optinal cmmand a pharmacphre byfeatures pressing (Cmd-F9 n [2]. OS Perfrm X) []. screening, examine resulting hits, and inject mst suitable nes back int prtein binding site [3]. Reset three left features as mandatry using Pharmacphre -> Unmark Features as Wherecmmand, t g frm Optinal sere: right tw features as ptinal and repeat screening. Examine resulting hits, and inject mst Exprting a pharmacphre t Catalyst, Phase and MOE suitable nes back int prtein binding site. In Structure-based perspective, minimize each fragment and bserve ir Pharmacphre mdeling: Creating shared and merged feature pharmacphres behavir. Using viewer ptin in hierarchical view, make tw fragments visible that verlap with ne atm. Delete ne f Custmizing pharmacphre creatin preferences verlapping atms and merge mlecules using Mlecule -> Merge Visible Mlecules cmmand. Create a bnd between bth fragments. Minimize resulting new mlecule and create a pharmacphre mdel. Analyze interactins and cmpare m t mdel generated frm initial ligand. Repeat this prcedure fr r fragment cmbinatins []. 1 Where t g frm here: 2 3 Re-dck, minimize, and re-scre newly created mlecules in rder t identify mst interesting candidates Exprt mst favrable designs as an Excel spreadsheet, r as 3D r 2D sdf files

11 CARDCARD 11 Creating a pharmacphre frm a single prtein-ligand cmplex Finding druggable pckets in prteins with LigandScut advanced basic 10 minutes minutes Macrmlecular view Macrmlecule view Active site view Active site view User Cntrls User Cntrls Advanced cntrls (pt.) Advanced cntrls (pt.) Dwnlad PDB file using Pcket finder widget Adjust parameters fr Dwnlad PDB file using Ligand bx Change bnd type -letter cde Ligand bx buriedness and threshld -letter cde Create pharmacphre Change atm type Perfrm pcket analysis Create multiple Discver crrect (buttn r menu) Create newbinding bnd Select and create a new pckets ligand OR: Create pharmacph Mve t cre Fcus n active site and re fr Catalyst/MOE/ Mve t envirnment binding pcket Perfrm dcking switch t active site view Phase Cmpund Perfrm ap-site switching Check and eventually crpharmacphre mdel rect ligand chemistry generatin Create a single-click phar- macphre Descriptin: Descriptin: Type 1ke6 in upper right area f screen, an press buttn Dwnlad 1ke6 [1]. The prtein will be dwnladed and displayed macrmlecule [2]. Click n Dwnlad [1]. yellw bx within representing ligand -->an Type eq3 in (this upperisright area f screenview ) and press buttn Frm prtein drpdwn menu, select Mlecule animated zm begins ending in [3]. Since ligand structures in PDB nly cntain incmplete infrmatin, yu shuld Calculate Pckets [2]. A grid is calculated ver entire prtein structure and grid pints are evaluated accrding t ir buriedness always check wher all bnds are typed crrectly. Bnd types can be changed by selecting a bnd (by clicking n it eir in and ir number f neighbring grid pints. Iscntur surfaces are generated [3]: Orange clred issurfaces are cnsidered t 2D r 3D view) and by using retype bnd buttn [] r keys 1, 2, r 3. Once ligand is chemically crrect, create highlight druggable pckets, where Ctrl-F9 dark grey clredn nes cnsidered t small t accmmdate a drug-like ligand. Values fr a pharmacphre by pressing (Cmd-F9 OSare X) []. buriedness and threshld can be mdified by mving sliders n Pcket-GRID widget [3], while clicking n Create pckets Where g frm buttn willt define a newhere: binding site highlighted by yellw bx []. Clicking n yellw bx will zm int Active site view Exprting a pharmacphre t Catalyst, Phase and MOE and frm here, dcking r ap-site pharmacphre creatin can be perfrmed. Pharmacphre mdeling: Creating shared and merged feature pharmacphres Custmizing pharmacphre creatin preferences Where t g frm here: Dck ligands int newly defined binding site Create ap-site pharmacphre mdels 1 2 3

12 CARDCARD 12 Creating a pharmacphre single prtein-ligand cmplex Creating pharmacphre mdels infrm emptya binding sites advanced basic 2 minutes minutes Macrmlecule view Macrmlecule view Active site view Active site view Hierarchical view User Cntrls User Cntrls Advanced cntrls (pt.) Advanced cntrls (pt.) Open PDB file Pcket finder widget Perfrm virtual screening Dwnlad PDB file using Ligand bx Change bnd type Find ptential binding Ligand bx Refine pharmacphre -letter cde Create pharmacphre Change atm type pcket Ligand accessible surface mdels frmnew retrieved Discver crrect (buttn r menu) Create bndhit Switch t active site view prperty display mlecules ligand OR: Create pharmacph Mve t cre Fcus n active reap frsite Catalyst/MOE/ Mve t envirnment Create ap site grids site and Define Mve pharmacphre switch t active site view pharmacphre Phase Cmpund switching Create ap site pharmacsettings features in 3D space Check and eventually crphre mdel rect ligand chemistry Create a single-click phar- macphre Descriptin: Descriptin: Type 1ke6 in upper screen, an press buttn Dwnlad 1ke6 The prtein will be dwnladed Type ek3 in upper rightright area area f fscreen, and press buttn Dwnlad ek3 [1]. The[1]. prtein will be dwnladed and and displayed (this is macrmlecule view ) [2]. Click n yellw bx within prtein representing ligand - an displayed in macrmlecule view. Search fr druggable pcket using LigandScut Pcket Finder (see tutrial card 11) and animated zm begins ending in [3]. Since ligand structures in PDB nly cntain incmplete infrmatin, yu shuld define binding site. Zm int it by clicking n yellw bx. Calculate entire pssible pharmacphre interactins by always check wher all bnds are typed crrectly. Bnd types can be changed by selecting a bnd (by clicking n it eir in selecting Pharmacphre -> Create Ap Site Grids [2]. Mdify ap site pharmacphre feature prbabilities by mving sliders 2D r 3D view) and by using retype bnd buttn [] r keys 1, 2, r 3. Once ligand is chemically crrect, create in Ap-Site Pharmacphre widget []: Mving right will make features less prbable, mving m t left will a pharmacphre by pressing Ctrl-F9 (Cmd-F9sliders n OStX) []. make features mre prbable. Restre default settings by clicking return back symbl lcated at right f slider. By clicking t g frm here: Where Create buttn, Ap-Pharmacphre Settings dialgue will appear []. Create a mdel by selecting nature and number f Exprting a pharmacphre t Catalyst, Phase and MOE features that yu want t be cnsidered in autmated generatin prcess. Start with HBA, HBD, and hydrphbic features. Repeat Pharmacphre mdeling: Creating shared and merged feature pharmacphres prcedure fr psitive and negative inizable features. Refine mdel by mdifying final number f features, merging sme Custmizing pharmacphre creatin preferences f m, r mving m t new psitins. Select grid pints and add features if necessary. Add r remve exclusin vlume spheres. Use mdel fr virtual screening r fr re-scring dcked pses. Where t g frm here: 1 2 Insert hits btained frm VS int binding site 3 Create new pharmacphre mdels frm minimized hit structures btained Re-scre pses btained frm mlecular dcking

13 CARDCARD 13 Creating a pharmacphre frm a single prtein-ligand cmplex Dcking a mlecule int a binding site using built-in AutDck prgram intermediate basic 2 minutes minutes Macrmlecule view Macrmlecule view Active site view Active site view Table view User Cntrls User Cntrls Advanced cntrls (pt.) Advanced cntrls (pt.) Dwnlad PDB file using Ligand bx Filter pses based n Dwnlad PDB file using Ligand bx Change bnd type letter cde Insert smiles string pharmacphre features -letter cde Create pharmacphre Change atm type Switch t active site view Dck(buttn mlecule Minimize pses Discver crrect r menu) Create new bnd Insertligand mlecule int Analyze dcking pses Re-scre pses by estimating OR: Create pharmacph Mve t cre Fcus re fr Catalyst/MOE/ free Mve t envirnment binding siten active site and energy f binding switch active site view Phase Cmpund switching Perfrm t dcking Create new binding site Check and eventually crexperiment Dck mlecules int new rect ligand chemistry binding site Create a single-click phar- macphre Descriptin: Descriptin: Type 1ke6 in upper right area f screen, an press buttn Dwnlad 1ke6 [1]. The prtein will be dwnladed and displayed (this isright macrmlecule [2]. Click n Dwnlad yellw bx1ke7 within representing ligand Type 1ke7 in upper area f screen,view ) and press buttn [1]. The prtein prtein will be dwnladed and - an animated zm begins ending in [3]. Since ligand structures in PDB nly cntain incmplete infrmatin, yu shuld displayed in macrmlecule view. Click n yellw bx within prtein representing ligand in rder t zm int always check wher all bnds are typed crrectly. Bnd types can be changed by selecting a bnd (by clicking n it eir in binding site. Insert mlecules as sdf that yu want t dck int binding site, using File -> Insert [2] r by pasting a smiles file 2D r 3D view) and by using retype bnd buttn [] r keys 1, 2, r 3. Once ligand is chemically crrect, create (Cntrl-V ). Select mlecules that Ctrl-F9 yu want t dckn frm table belw 3D windw. Prceed with dcking experiment by a pharmacphre by pressing (Cmd-F9 OS X) []. using Mlecule -> Dck Ligands -> AutDck.2 r AutDck Vina 1.1 [3]. After experiment is finished, results will be displayed Where g [], frm here: in tablet view each cmpund ranked accrding t mst favrable interactin energy. Analyze dcked pses by creating Exprting a pharmacphre t Catalyst, Phase and MOE pharmacphre mdels fr viewing interactins with prtein. Filter pses based n interactins cmmn t thse f Pharmacphre mdeling: Creating shared and merged feature pharmacphres riginal ligand. Custmizing pharmacphre creatin preferences Where t g frm here: Minimize dcked pses Calculate free energy f binding and determine MMFF9 estimated binding enthalpies Calculate pharmacphre fit scre based n mdel generated frm riginal ligand structure Create a new binding site by selecting amin acids with muse (hld left buttn while pressing Shift key, mve ver regin yu want t investigate and create a binding site frm menu Mlecule -> Create New Binding Site), r use pcket finder (see tutrial card 11) fr finding a new druggable pcket and repeat dcking experiment.

14 CARDCARD 1 Creating a pharmacphre frm a single prtein-ligand cmplex Analyzing Mlecular Dynamics Trajectries with LigandScut advanced basic 1 minutes minutes Macrmlecule view Macrmlecule view Active site view Active site view User Cntrls User Cntrls Advanced cntrls (pt.) Advanced cntrls (pt.) Open PDB file Ligand bx Create shared feature Dwnlad PDB file using Ligand bx Change bnd type Switch t active site view Mlecular dynamics mdels -letter cde Create pharmacphre Change atm type Lad mlecular dynamics trajectry Widget Perfrm virtual Discver crrect (buttn r menu) Create newscreening bnd trajectry file Playback trajectry with MD derived ligand OR: Create pharmacph Mve t cre Fcus active site and Create repharmacphre fr Catalyst/MOE/ Mve t envirnment Analyze n trajectry pharmacphre mdels switch t active site view mdels Phase Cmpund switching Create pharmacphre Check and eventually crmdels Align pharmacphre rect ligand chemistry Create a single-click phar- mdels macphre Descriptin: Descriptin: Type 1ke6 in upper right area f screen, an press buttn Dwnlad 1ke6 [1]. The prtein will be dwnladed and (this is macrmlecule [2]. Click dynamics n yellw bx within prtein ligand - an Lad displayed PDB file cntaining initial frame f view ) yur mlecular simulatin (1ke7_start.pdb) [1].representing Click n yellw bx within animated zm begins ending in [3]. Since ligand structures in PDB nly cntain incmplete infrmatin, yu shuld prtein representing ligand in rder t zm int binding site. Lad DCD file f mlecular dynamics trajectry always check wher all bnds are typed crrectly. Bnd types can be changed by selecting a bnd (by clicking n it eir in (1ke7_trajectries.dcd) [2] and select trajectry in hierarchical view [3]. Press play buttn in MD trajectry widget [] fr 2D r 3D view) and by using retype bnd buttn [] r keys 1, 2, r 3. Once ligand is chemically crrect, create viewing results f simulatins []. Pause a pharmacphre by dynamics pressing Ctrl-F9 (Cmd-F9 n OSatX)any [].time and generate a pharmacphre mdel. Mve pharmacphre mdel int alignment perspective. Repeat this several times. In alignment perspective, analyze features t g frm here: [6]. thatwhere are cmmn in all mdels Exprting a pharmacphre t Catalyst, Phase and MOE Pharmacphre mdeling: Creating shared and merged feature pharmacphres Where t g frm here: Custmizing pharmacphre creatin preferences Align pharmacphre mdels generated frm MD trajectry Create shared feature and merged feature mdels frm different pharmacphre families Perfrm virtual screening with different mdels 1 2 3