FLAP

Introduction

FLAP is used to refine results obtained from an analysis performed with PocketQuery/ZINCPharm online tools

PocketQuery is a web service for interactively exploring not only hot spot and anchor residues, but hot regions, defined by clusters of residues, at the interface of protein-protein interactions.

Instructions

Pocket Query

Open a browser and connect to http://pocketquery.csb.pitt.edu/

Click on Search

Type PDB code

• You can Add Search Criteria
• Chain to select the chain under study

Click on Search to submit the query

Analyze clusters to find pockets using the two panels Residues and Export

• Pay attention to the correct definition of the ligand protein and the receptor protein

When the pocket is found, it is possible to direclty export, panel Export, in ZINCPharmer (Send to ZINCPharmer)

• More than one cluster could be used to define the fine pocket

ZINCPharmer

Open the browser and connect to http://zincpharmer.csb.pitt.edu/

• If you sent the pocket to ZINCPharmer from PocketQuery the browser is already open
• Java could have problems, but only for visualization of surfaces thus it is possible to proceed

Use the panel Pharmacophore to define the hot spots that will be used in the screening

Click on Submit Query

After the screening, Save Results to obtain a file sdf with the coordinates of compounds that better respond to the pharmacophore criteria

• The coordinates refer to the conformation that is able to optimize interaction with the receptor on the basis of ZINCPharmer force field
• It is possible to enable or disable different pharmacophoric points (HBA, HBD, Hydrophobic points and Aromatic)
• Probably, it should be convenient to try different pharmacophores and save the corresponding results, all the results can be merged to obtain the final dataset of candidates to submit to FLAP

FLAP refinement

Launch FLAP

Database creation

Database | Create -> FLAP open the FLAP Database Creation window

• Create e directory and use it as the Database location

Create Database

• Select options
• We do not have the protonation tool thus the molecules have to be in the correct ionization state
• Select do fully energy minimization with default parameter (25 kcal/mol)
• Duplicate refuse

Start import

Setting of the target

Structure-based | Load protein

• Fix pdb
• Select how water molecules will be considered

Ok

FLAP open the Target setup window

Pockets panel

• Option Search for pockets can be used to search automatically the pockets
• Option Search by residue can be used to focus the search a region around one or more residuals
• Select one or more residue by picking with the mouse in the Pocket by residue window, after picking the mouse a residue line with all residue

Select pocket

• Give a name to the pocket in Pocket name
• OK

Structure-Based Virtual Screening (SBVS)

In the Database panel select molecules, if no molecules are selected the screening is carried out on the entire database

• Set the Screening name
• Choose the target: add target and select the pocket from the Define pocket and Enable Objects window
• Parameters
• select pose optimization on shape
• The option onGlobSum with highest accuracy crashed FLAP but more investigations are required to understand the problem
• Accuracy level 2 (4=High, 1=Low) successfully tested with a dataset of about 3000 molecules in a reasonable time

Finally, analyze the results in the Screening panel