MM-GBSA Residue Scanning - Homology Suggestions Dialog Box

Identify candidate residues for mutation, using homology modeling or structural criteria. Homology modeling can be applied to select residues by variability or conservation, structural proximity or properties. Structural criteria can be applied for solvent accessible surface area and interactions (contacts) with the protein or a selected molecule.

To open this panel, click the Tasks button and browse to Biologics → MM-GBSA Residue Scanning Calculations.

  • Using
  • Features
  • Additional Resources

Using the Homology Suggestions Dialog Box

The main tasks in this panel are to create a homology model for one of the protein chains, select residues by applying criteria based on the homologs, optionally select criteria based on the 3D structure of the protein, then apply the selection to the residue table in the Residue Scanning panel by clicking Save.

The homologs used in the model can be imported (click Import homologs) or located by running a BLAST search. The homologs must then be aligned, which you can do manually or by running an alignment (click Align Homologs or the toolbar alignment buttons).

If you don’t have homologs for your protein, you can run a BLAST search to find homologs.

  1. Click Run a Blast Search.

    The BLAST Search Settings dialog box opens, so you can change settings if you want.

  2. Click Start Job in this dialog box, after changing any settings.

    The job starts, and its log file is displayed in the Job Progress dialog box.

At the end of the job, the BLAST Search Results dialog box opens, so that you can choose how many of the homologs you want to use. You can do one of the following to select homologs:

  • Select the homologs in the table (with shift-click and control-click).

  • Enter the number of the top homologs in the text box at the bottom of the panel and click Select Top.

When you have selected the homologs, click Incorporate Selected Rows to add the structures to the sequence viewer.

Homology Suggestions Dialog Box Features

Chain option menu

Select the chain to show in the sequence viewer and use for generating homology suggestions.

Show antibody annotation options option

Show options relating to antibody annotation. When this option is selected, the antibody hypervariable loops are annotated in the sequence viewer when you select the L or H chain of an antibody. This option also displays the Antibody numbering scheme option menu, which affects the annotation.

Upper toolbar

This toolbar has tools for importing sequences, undoing and redoing sequence editing, and finding patterns in a sequence.

  • Import homologs button—Import sequences into the sequence viewer to use for the homology criteria. Only the sequences are needed, no structural information is required. The choices are:

    • From File—Open a file browser in which you can navigate to the desired location and select the file that contains the sequence. If you are not using the sequences
    • From Workspace—Import the sequence for the structure that is displayed in the Workspace.
    • From PDB ID—Import the sequence from the specified PDB ID. Opens the Enter PDB ID dialog box, in which you can enter the PDB ID of the sequence. The sequence is retrieved from a local copy of the PDB if it is available, or from the RCSB web site, depending on the preference set for PDB retrieval.

  • Undo and Redo buttons—Undo or redo the actions taken in the sequence viewer.

  • Find Pattern text box—Find a particular pattern in a sequence. You can use an extended PROSITE syntax, which makes use of secondary structure and property information; the syntax is explained in the tooltip. See the Multiple Sequence Viewer/Editor — Toolbar topic for more information.

  • Arrow buttons—These buttons can be used to step through instances of the pattern, in the forward or reverse direction.

Sequence viewer

The sequence viewer displays the sequence of the Workspace structure and any sequences that you import or obtain from a search for homologs. You can drag a residue to align it and the residues downstream (to the right). Downstream gaps are preserved. Gaps can be locked or unlocked with the toolbar buttons below the sequence viewer, so that they are not filled when you drag to the left.

Alignment toolbar

This toolbar allows you to perform tasks related to sequence alignment, which can be done manually or automatically.

Lock gaps
Lock gaps in the sequences so that they are not filled when performing manual alignment. The locking is applied once. Gaps created after locking is done are not automatically locked. To lock them, click this button again. Locked gaps are indicated by a dash (-); unlocked gaps are indicated by a tilde (~). If you have a residue selection, gaps are only locked in the selected region.
Unlock gaps
Unlock gaps in the sequences after they have been locked. This allows gaps to be filled when performing manual alignment.
Pairwise Alignment
Align the selected sequences with the query sequence one at a time (pairwise), using ClustalW.
Multiple Alignment
Align the selected sequences simultaneously, using ClustalW. The sequence alignment is restricted to the selected columns in the sequence viewer. The alignment adds gaps in the sequence viewer as necessary.
Antibody numbering scheme option menu

Choose a numbering scheme for the residues in an antibody. This choice affects the annotation scheme that is used. This menu is only displayed if Show antibody annotation options is selected.

Run a Blast Search button

Run a Blast search to identify homologs to the query (parent) sequence. Opens the Blast Search Settings dialog box, in which you can make any settings for the search, then start the job.

Align Homologs button

Perform a multiple sequence alignment of the homologs to the query (parent).

Selection by homology criteria options

Set criteria for the selection of residues to mutate based on the homologs found. The variations found between the homologs and the query are used as a basis for choosing default mutations. Each of the criteria that you select is applied, so the residues must meet all specified criteria.

  • Variability at position >/< N %—Filter residues based on the percentage variability at the residue position. The percentage variability is the normalized Shannon entropy for residue variation at the position, expressed as a percentage. Choose whether to apply a minimum or a maximum variability from the option menu, and specify the percentage threshold in the text box.

  • Variability at position >/< N residue types—Filter residues based on the variation in the residue type at the residue position. The variability is the number of residue types observed at the position. Choose whether to apply a minimum or a maximum variability from the option menu, and specify the threshold for the number of residue types in the text box.

  • Ignore positions with group conservations—Ignore (do not select) residues that are strongly conserved or that are either strongly or weakly conserved. Select the appropriate option for strong conservation or both strong and weak conservation. The definitions of strong and weak conservation are those used by ClustalW:
    Strongly conserved groups: STA, NEQK, NHQK, NDEQ, QHRK, MILV, MILF, HY, FYW.
    Weakly conserved groups: CSA, ATV, SAG, STNK, STPA, SGND, SNDEQK, NDEQHK, NEQHRK, FVLIM, HFY.

  • Ignore parent sequence in applying above criteria—Ignore the parent (query) sequence when applying the variability or conservation criteria.

  • Parent residue different than N % homologs at position—Select residues for which the parent residue is different from more than the specified percentage of homologs at the residue position.

Parent structure 3d criteria options

Set criteria for selection of residues based on the 3D structure of the parent (the structure that was analyzed). The criteria can be based on solvent-accessible surface area (SASA) and interactions between residues. Interactions are determined by a distance cutoff: any residue that has atoms within 4 Å of the side chain is considered to interact.

  • Solvent accessible surface area—Select this option to filter residues by their solvent-accessible surface area (SASA) relative to an isolated residue of the same type, and set a threshold for the maximum or minimum allowed relative SASA. This option is useful for locating surface residues. (The SASA is calculated by the Lee-Richards method [12], which is also used in Canvas and Phase).

  • Residue side chain makes no more than N interactions with protein—Select this option to filter out residues whose side chains interact with the protein, and set the maximum number of such interactions.

  • Residue side chain interacts/does not interact with molecule N—Select this option to filter residues by their interaction with a selected molecule. Choose whether to allow or disallow the interaction from the option menu, and specify the molecule in the text box, or select Pick and pick the molecule in the Workspace.

If you select residues only by their structural attributes, there are no suggestions for the mutations, so the result is that the matching residues are selected in the residues table. You can then set the mutations for these residues. If structural attribute criteria are combined with homology criteria, then the mutations are set as well as selecting the residues.

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