Protein Surface Analyzer Panel

Analyze a protein surface into patches that represent distinct types of interactions: positively charged, negatively charged, and hydrophobic. For each patch a range of properties is calculated, to characterize the interaction and its intensity. These patches can be used to identify possible interaction hotspots or propensities to interact.

To open this panel, click the Tasks button and browse to Biologics → Protein Surface Analyzer.
To open this panel from the entry group for the results of an antibody structure prediction job, use the Workflow Action Menu .

Overview
Using
Features
Additional Resources

Overview of the Protein Surface Analyzer Panel

Developability assessment is a key stage of the lead optimization stage of the protein-based therapeutic development process. One property of particular interest during developability assessment is aggregation propensity as it is a key concern during both the production and storage of protein-based therapeutics, especially in regards to protein stability[1]. Protein aggregation is of the most common and most difficult issues encountered in the developability assessment stage [1,2,3].

As aggregation propensity has historically been expensive to predict experimentally, there is a clear need for computational tools that could help predict the relative aggregation propensity of a set of protein variants. AggScore, a structure-based descriptor, is one such tool that can be used to prioritize protein variants based on predicted aggregation propensity. See the AggScore: Prediction of aggregation-prone regions in proteins based on the distribution of surface patches for more information[1].

While hydrophobic patches have traditionally been viewed as a key driver of aggregation, AggScore considered whether those patches are surrounded (in 3-dimensional space) by neighboring polar patches. Residue-specific contributions to aggregation (as determined by positive, negative, and hydrophobic patches) are calculated on the interaction surface of the protein and smoothed over a window of five residues. This smoothing allows for the polar patches to limit or possibly negate entirely the predicted impact a hydrophobic patch could have on aggregation propensity[1].

Consider the two 5-residue peptides below. While both have polar phenylalanine in the center, the phenylalanine in the peptide on the left is neighbored by two polar serines, while the phenylalanine in the peptide on the right is surrounded by alanines.

AggScore results for two 5-residue peptides

As the phenylalanine in the peptide on the left is flanked by two polar residues which counteract the impact the hydrophobic patch could have on aggregation propensity, the AggScore for all of the residues is zero. For the peptide on the right, however, we can see that the AggScore associated with the phenylalanine is 3.2, since it is surrounded by non-polar alanine residues which do not negate the impact of the hydrophobic patch. It is the interplay between the polar and hydrophobic patches in a 3-dimensional environment that afford the AggScore a predictive power beyond that of the commonly used sequence-based predictors.

The AggScore function provides propensity values for each amino acid position in a protein, thus allowing for the prediction of aggregation hotspots within the protein. It also provides a single value for an entire protein, which can be used for rank-ordering several variants by their relative aggregation propensity. It is important to note that AggScore is not dependent on the presence of natural amino acids and is not trained on beta-amyloid aggregation data.

References:

[1] Sankar K, Krystek Jr SR, Carl SM, Day T, Maier JKX. AggScore: Prediction of aggregation-prone regions in proteins based on the distribution of surface patches. Proteins. 2018; 86: 1147–1156. https://doi.org/10.1002/prot.25594

[2] Redington M. Jennifer , Breydo Leonid and Uversky N. Vladimir, When Good Goes Awry: The Aggregation of Protein Therapeutics, Protein & Peptide Letters 2017; 24 (4). https://dx.doi.org/10.2174/0929866524666170209153421

[3] “Aggregation prediction with protein surface analyzer.” 2024, https://newsite.schrodinger.com/life-science/learn/white-papers/aggregation-prediction-protein-surface-analyzer/

Using the Protein Surface Analyzer Panel

The purpose of the panel is to allow an evaluation of the possible regions ("patches") on the protein surface that might contribute to aggregation, at a level higher than individual residues. As protein aggregation is a complex issue, the quantitative measures given here are intended for guidance, not for prediction.

The protein on which the analysis is done should be prepared with the Protein Preparation Workflow Panel first.

Ligands and other hetero groups can be included in or excluded from the analysis. By default, covalently bound ligands are included, and other groups are excluded. However, not all hetero groups might be considered as "ligands", as the ligands are detected using Maestro's ligand detection settings. These settings can be changed in the Ligand Detection Preferences settings in the Preferences Panel. The choice of ligands to include or exclude is made in the Protein Surface Analyzer - Settings Dialog Box.

For example, the covalently-bound sugars in 5DK3 appear in the hetero group table of the Settings dialog box and are included by default if you allow the relevant residues (NAG, BMA, MAN, and FUC) to be detected as ligands in the Ligand Detection Preferences of the Preferences Panel and you increase the maximum heavy atom count for ligand detection to cover these groups. If you make these changes after opening the Settings dialog box, you should close it and reopen it to see the updated list of hetero groups.

If you want to share the results with someone else, you can save the entire analysis including the structure and surface with the Save Analysis button, then send them the .psazip file, which they can then load in Maestro with the Load Analysis button.

Once you have located potential aggregation regions on a protein, you might want to mutate residues in these regions to reduce the tendency to aggregate. One way of doing this is to perform a set of mutations to reduce the size of the aggregation regions. You can create a set of structures that have single mutations at selected sites, which you choose from the residues in the aggregation regions, as follows:

Select residues in the Workspace that you have identified as likely to promote aggregation.

Click the Tasks button and browse to Biologics → MM-GBSA Residue Scanning Calculations to open the MM-GBSA Residue Scanning Panel.

Choose Workspace (selected residues only) from the Import structure from option menu.

Click Import.

The residues that you identified as contributing to aggregation are listed in the table in the Residues tab. You can then select any of them for mutation, define the mutations, and run the job. See MM-GBSA Residue Scanning Panel for details of setting up a residue scanning job.

Another option is to mutate a single residue or a loop. Mutating a loop (“loop swap”) is useful if you want to mutate more than one residue and the residues are adjacent. For this purpose, you can use the Residue and Loop Mutation Panel. The loop to change is defined by selecting the residues in the Workspace.

To run protein surface analysis from the command line, you can use the following command. Run the command with -h for more information.

$SCHRODINGER/run -FROM psp protein_patch_calculation.py

For information on command options, see protein_patch_calculation.py Command Help.

Use structure from menu and button

Select the source of the structure to analyze. The button that appears to the right depends on the choice of source.

Workspace—Use the structure from the Workspace. The button to the right is labeled Analyze; when you click it, the protein is analyzed.
Workspace (selected residues only)—Use the selected residues from the structure in the Workspace. The button to the right is labeled Analyze; when you click it, the protein is analyzed.
File— Use the first structure from the specified Maestro file. The button to the right is labeled Import and Analyze. When you click it, a file selector opens so that you can choose the file. Once it is read, the protein is analyzed.

If the protein is not properly prepared, you are prompted to prepare it with the Protein Preparation Workflow Panel.

Settings button

Make settings for the minimum area of the positive, negative, and hydrophobic patches, set a threshold value for each that determines the extent of the patch, and set the color and transparency for the patches. Also select heterogroups for inclusion as part of the surface analysis. Opens the Protein Surface Analyzer - Settings Dialog Box.

Patch Browser tab

This tab contains a table that lists all the patches, giving the type (pos, neg, hyd), the patch size, the patch energy score, and the patch intensity (score/size).

For each patch there is also a Details button, which opens a dialog box that lists the residues in the patch and gives information on surface exposed residues and reactive sites, as well as reproducing the information in the table. You can export this information to a CSV file by clicking the Export button in the dialog box.

You can select rows in the table with the usual click, shift-click, and control-click actions. Right-clicking on a table cell displays a menu, from which you can choose from the following actions:

Show in Workspace—Show this patch in the Workspace, along with any other patches.
Show Only Selected in Workspace—Show only the selected patches in the Workspace, and hide all others.
Hide in Workspace—Hide this patch in the Workspace, leaving any other patches displayed.
Details—Open the Details dialog box to show detailed information on the patch (see below)
Export Details—Export the details for the patch to a CSV file.
Select All—Select all patches.
Invert Selection—Invert the selection of patches: selected patches are deselected, unselected patches are selected.
Delete—Delete the selected patches from the table.

Aggregation tab

This tab contains a table that lists the residues contributing to each patch, along with various properties that relate to the propensity to agggregate. Some of the scores are available as atom properties for the Cα atoms, which you can display as labels (see Labeling Atoms and Bonds).

Residue Aggregation Profiles table

When you select a table row, the residue is selected in the Workspace, and the view zooms in on the residue listed in that row if you have Fit on select selected. You can select multiple residues (rows) to send to the Residue Scanning panel, which you do by clicking Export to Residue Scanning. The columns are described below.

Residue	Chain name, residue name, and residue number
Topo	Topology of the residue if it is in an antibody. The particular region of the antibody in which this residue is found (H1, H2, H3, etc) is shown in this column, which is only present if the protein is an antibody.
Patch	Index of the patch to which the residue contributes
Size	Size of the patch in Å²
Patch Score	Patch energy score (sum of potential energies from all points on the patch).
Contribution	Contribution of the residue to the energy score of the patch, color coded by magnitude and type (positive: blue, negative: red, hydrophobic: green).
AggScore	Composite aggregation propensity score, based on surface patch projections of hydrophobic and electrostatic atoms, factoring in hydrophobicity and charge propensities. Only positive aggregation propensity is considered. The global AggScore is added to the structure as a Maestro property.
Zyggregator	Zyggregator score [15]. This score is based on sequence, folding propensities, and charge, parameterized using short peptides and validated with a number of known amyloidogenic peptides, validated on 24 fibril-forming peptides.
Aggrescan	Aggrescan score [14]. This is a sequence-based prediction method based on aggregation propensity data of point mutations of beta-amyloid in region 17-21.
ASA	Percentage of the residue's solvent-accessible surface area compared to that of the residue when in the central position of a reference trimer.

Note on SASA values: There is no consensus on the ideal SASA cut-off value. A suitable cut-off depends on the goal of the analysis. A small cutoff (10%-20%) may be appropriate if the goal is to identify any residues that are likely to participate in interfacial interactions whereas a >50% cut-off has been recommended in some studies [19] intended to identify residues that pose significant aggregation risk.

Profiles button

Show bar plots of the contributions, scores, and solvent-accessible surface area on a per-residue basis. Opens the Aggregation Profiles Panel.

Export to Residue Scanning button

Export the residues that are selected in the table to the MM-GBSA Residue Scanning Panel. This panel opens with these residues selected for mutation.

Reactive Residues tab

This tab lists reactive surface-exposed residues by patch. As aggregation can be promoted by covalent bonding, the residues listed here should be considered as potential sites for aggregation.

Residue and site selection options: Select the residues or the sites to show in the table. All residues or sites of the selected type are shown. You can choose from the four residue types Arg, Cys, Trp, Tyr; disulfide bonds; or deamidation, glycosylation, proteolysis, or oxidation sites (as defined in the Reactive Protein Residues panel).
Minimum side chain accessibility text box: Set the minimum solvent accessibility of the side chains of the residues for the residues to be listed in the table. Residues that have a smaller solvent accessibility of the side chain are not shown.
Residue table: This table lists the residues, giving the patch number, the side chain accessibility (ASA), and a check mark in the appropriate remaining column if the residue contributes to a disulfide bond or one of the four types of reactive sites. When you select a table row, the Workspace view zooms in on the residue listed in that row. You can select multiple residues (rows) to send to the Residue Scanning panel.
Export to Residue Scanning button: Export the residues that are selected in the table to the MM-GBSA Residue Scanning Panel. This panel opens with these residues selected for mutation.

Properties tab

This tab displays some general protein properties, including sums of various surface areas and the sum of the AggScore values.

Options

The options below the tabs allow you to control the patches that are displayed and relevant Workspace behavior or appearance.

List patch type options

Select the types of patches to display in the table.

Positive—Show hydrophilic patches that have a positive potential.
Negative—Show hydrophilic patches that have a negative potential.
Hydrophobic—Show hydrophobic patches.
Only selected patches—Show only the patches for the residues that are selected in the Workspace.

Show Workspace labels option

Label the patches with the patch number from the table. The labels are colored with the patch type.

Color by option menu

Color the protein surface according to the type of patch, or by the AggScore values. The coloring is also applied to certain columns in the tables in the Patch Browser and Aggregation tabs.

Save Workspace Selection button and menu

Save the Workspace selection to a named location, which can be retrieved in other tabs of the panel. The Workspace selection is made by selecting one or more rows in any of the tabs. A dialog box opens so you can name the selection. This process essentially saves the residues in the patches. You can retrieve the selection from the menu, and it is applied to the Workspace. If you retrieve a selection in a tab that works with patches, all the residues in the patch are selected. If a residue is in more than one patch, all patches it is in are selected.

Fit on select option

When a patch is selected in the table, center and zoom in to the patch residues in the Workspace.

Load Analysis and Save Analysis buttons

Load and save surface analysis data. The entire analysis is saved, including the structure, surface, data, and panel settings, in a zip file with a .psazip extension. When an analysis is loaded, the structure and surface are imported and the panel and display are set to the state they were in when the analysis was saved. As the file contains all relevant data, it can be opened by anyone in a Maestro session.

Export button

Export the patch analysis data to a CSV file. Opens the Export Data dialog box, in which you can select the tables (Patch Browser, Aggregation, Reactive Residues) to export, then opens a file selector so you can name the file. The three tables have the extensions -patches.csv, -aggregation.csv, and -reactive.csv.

Reset button

Clear the panel of all data and reset it to the default state.

Tutorials