AppendAssemblyMover

AppendAssemblyMover is a derived AssemblyMover that builds a SEWING assembly from a predetermined starting structure.

Table of Contents

Description

AppendAssemblyMover is used to incorporate a functional element of a native structure into a larger backbone. It is most commonly used to design binding partners for a protein using a binding peptide as a starting structure. It has also been used to design protein backbones around a native ligand binding site.

Usage

NOTE: This is currently out of date but will be automatically updated when the new version of SEWING is merged.

Autogenerated Tag Syntax Documentation:

Builds an assembly around the segment provided in the input PDB file

<AppendAssemblyMover name="(&string;)" start_temperature="(0.6 &real;)"
        end_temperature="(0.6 &real;)" add_probability="(0.05 &real;)"
        delete_probability="(0.005 &real;)"
        conformer_switch_probability="(0 &real;)"
        window_width="(4 &positive_integer;)"
        minimum_cycles="(10000 &non_negative_integer;)"
        maximum_cycles="(100000 &non_negative_integer;)"
        model_file_name="(&string;)" hashed="(false &bool;)"
        edge_file_name="(&string;)" max_segments="(100 &non_negative_integer;)"
        max_segment_length="(100 &non_negative_integer;)"
        output_pose_per_move="(false &bool;)"
        recover_lowest_assembly="(true &bool;)"
        recursive_depth="(1 &non_negative_integer;)"
        pose_segment_starts="(&int_cslist;)" pose_segment_ends="(&int_cslist;)"
        pose_segment_dssp="(&string;)" strict_dssp_changes="(false &bool;)"
        set_segments_from_dssp="(false &bool;)" match_segments="(&int_cslist;)"
        partner_pdb="(&string;)"
        required_resnums="(&refpose_enabled_residue_number_cslist;)"
        max_recursion="(1 &non_negative_integer;)"
        modifiable_terminus="(B &string;)" output_partner="(true &bool;)"
        extend_mode="(false &bool;)" start_node_vital_segments="(all &string;)"
        required_selector="(&string;)" >
    <AssemblyScorers >
        <InterModelMotifScorer name="(&string;)" weight="(1.0 &real;)" />
        <IntraDesignTerminusMotifScorer name="(&string;)" weight="(1.0 &real;)"
                optimum_distance="(&real;)" maximum_unpenalized_variance="(&real;)" />
        <LigandScorer name="(&string;)" weight="(1.0 &real;)"
                ligand_interaction_cutoff_distance="(5.0 &real;)" />
        <MotifScorer name="(&string;)" weight="(1.0 &real;)" />
        <PartnerMotifScorer name="(&string;)" weight="(1.0 &real;)" />
        <SegmentContactOrderScorer name="(&string;)" weight="(1.0 &real;)" />
        <StartingNodeMotifScorer name="(&string;)" weight="(1.0 &real;)" />
        <SubsetPartnerMotifScorer name="(&string;)" weight="(1.0 &real;)"
                region_start="(1 &non_negative_integer;)"
                region_end="(2 &non_negative_integer;)" />
        <TerminusMotifScorer name="(&string;)" weight="(1.0 &real;)"
                partner_residue="(&non_negative_integer;)" optimum_distance="(&real;)"
                maximum_unpenalized_variance="(&real;)" terminus="(&string;)" />
        <TopNMotifScorer name="(&string;)" weight="(1.0 &real;)"
                scores_to_keep="(1 &non_negative_integer;)" />
    </AssemblyScorers>
    <AssemblyRequirements >
        <ClashRequirement name="(&string;)"
                maximum_clashes_allowed="(0 &non_negative_integer;)"
                clash_radius="(4.0 &real;)" />
        <DsspSpecificLengthRequirement name="(&string;)" dssp_code="(X &dssp_enum;)"
                maximum_length="(100 &non_negative_integer;)"
                minimum_length="(0 &non_negative_integer;)" />
        <KeepLigandContactsRequirement name="(&string;)"
                contact_distance_cutoff="(2.5 &real;)" />
        <LengthInResiduesRequirement name="(&string;)"
                maximum_length="(10000 &non_negative_integer;)"
                minimum_length="(0 &non_negative_integer;)" />
        <LigandClashRequirement name="(&string;)"
                maximum_clashes_allowed="(0 &non_negative_integer;)"
                clash_radius="(4.0 &real;)" />
        <NonTerminalStartingSegmentRequirement />
        <SizeInSegmentsRequirement name="(&string;)"
                maximum_size="(10000 &non_negative_integer;)"
                minimum_size="(0 &non_negative_integer;)" />
    </AssemblyRequirements>
    <Ligands >
        <Ligand partner_ligand="(false &bool;)" pdb_conformers="(&string;)"
                alignment_atoms="(&string;)" auto_detect_contacts="(true &bool;)"
                ligand_resnum="(&refpose_enabled_residue_number;)"
                ligand_selector="(&string;)" >
            <Contact partner_contact="(false &bool;)"
                    contact_resnum="(&refpose_enabled_residue_number;)"
                    ligand_atom_name="(&string;)" contact_atom_name="(&string;)" />
            <Coordination coordination_files="(&string;)"
                    geometry_score_threshold="(1 &real;)" >
                <IdealContacts distance="(&real;)" angle="(109.5 &real;)"
                        dihedral_1="(30 &real;)" dihedral_2="(30 &real;)"
                        max_coordinating_atoms="(&non_negative_integer;)"
                        ligand_atom_name="(&string;)" />
            </Coordination>
        </Ligand>
    </Ligands>
</AppendAssemblyMover>

start_temperature: Temperature at start of simulated annealing
end_temperature: Temperature at end of simulated annealing
add_probability: Probability of adding a triplet of segments at any given step during assembly
delete_probability: Probability of deleting a terminal triplet of segments at any given step during assembly
conformer_switch_probability: Probability of switching ligand conformers during assembly. This should only be used if a ligand is present AND if you have provided conformers for that ligand.
window_width: Required number of overlapping residues for two segments to be considered a match. Used in hashless SEWING only (for hashed SEWING, this is determined by the hasher settings used when generating the edge file).
minimum_cycles: Minimum number of Monte Carlo cycles for assembly before completion requirements are checked.
maximum_cycles: Maximum number of Monte Carlo cycles for assembly before forced termination.
model_file_name: (REQUIRED) Path to file defining segments to use during assembly
hashed: Use the hasher during assembly to check overlap of all atoms? Requires an input edge file.
edge_file_name: Path to edge file to use during assembly (only used if hashed is set to true)
max_segments: Maximum number of segments to include in the final assembly
max_segment_length: Maximum number of residues to include in a segment
output_pose_per_move: Setting to true will output a pose after each move/revert.
recover_lowest_assembly: Setting to true will output the lowest assembly in the final pose
recursive_depth: How many nodes after the terminal node should we keep track of alignments for?
pose_segment_starts: Residue numbers of the first residue in each segment in the input pose
pose_segment_ends: Residue numbers of the last residue in each segment in the input pose. Length must match that of pose_segment_starts.
pose_segment_dssp: String indicating the secondary structure of user-specified segments, one character per segment (e.g. HLH for a helix-loop-helix motif). Length should match that of pose_segment_starts and pose_segment_ends if specified.
strict_dssp_changes: Segments require at least a 2-residue change in DSSP to specify a new segment
set_segments_from_dssp: Determine segment boundaries based on pose secondary structure
match_segments: Which segments from the input pose should we be able to append onto? Defaults to exterior segments.
partner_pdb: Name of PDB file containing binding partner for this assembly
required_resnums: Residue numbers of residues in the input structure that must be preserved
max_recursion: How many alignments from the end nodes should be stored in memory?
modifiable_terminus: Which terminus of the starting node may be modified.
output_partner: Should the output pdb contain the partner?
extend_mode: Should SEWING append only a single helix?
start_node_vital_segments: Which segments from starting node are vital? (terminal or all)
required_selector: Residue selector specifying residues in the input structure that must be preserved. The name of a previously declared residue selector or a logical expression of AND, NOT (!), OR, parentheses, and the names of previously declared residue selectors. Any capitalization of AND, NOT, and OR is accepted. An exclamation mark can be used instead of NOT. Boolean operators have their traditional priorities: NOT then AND then OR. For example, if selectors s1, s2, and s3 have been declared, you could write: 's1 or s2 and not s3' which would select a particular residue if that residue were selected by s1 or if it were selected by s2 but not by s3.

Subtag AssemblyScorers: The subtags of this tag define the AssemblyScoreFunction that will be used to evaluate assemblies

Subtag InterModelMotifScorer: Basic Motif score among non-adjacent helices

weight: How heavily will this term be weighted during scoring?

Subtag IntraDesignTerminusMotifScorer: Motif score to measure packing of assembly against partner PDB

weight: How heavily will this term be weighted during scoring?
optimum_distance: How far apart should that residue optimally be from the terminus?
maximum_unpenalized_variance: How far off from that can it be before it should be penalized?

Subtag LigandScorer: Scores how well ligand is buried based on orientation of nearby Ca's

weight: How heavily will this term be weighted during scoring?
ligand_interaction_cutoff_distance: The distance cutoff between ligand atom and c alpha that is considered an interaction.

Subtag MotifScorer: Basic Motif score among all helices

weight: How heavily will this term be weighted during scoring?

Subtag PartnerMotifScorer: Motif score to measure packing of assembly against partner PDB

weight: How heavily will this term be weighted during scoring?

Subtag SegmentContactOrderScorer: Favors assemblies whose segments form contacts with segments distant in the assembly

weight: How heavily will this term be weighted during scoring?

Subtag StartingNodeMotifScorer: Specifically scores packing against the starting node

weight: How heavily will this term be weighted during scoring?

Subtag SubsetPartnerMotifScorer: Motif score to measure packing of assembly against partner PDB

weight: How heavily will this term be weighted during scoring?
region_start: What is the first residue of the scored subset?
region_end: What is the last residue of the scored subset?

Subtag TerminusMotifScorer: Motif score to measure packing of assembly against partner PDB

weight: How heavily will this term be weighted during scoring?
partner_residue: Which residue of the partner should this scorer calculate distance to?
optimum_distance: How far apart should that residue optimally be from the terminus?
maximum_unpenalized_variance: How far off from that can it be before it should be penalized?
terminus: Which terminus should be scored?

Subtag TopNMotifScorer: Basic Motif score among all helices

weight: How heavily will this term be weighted during scoring?
scores_to_keep: How many scores from each pair should be counted?

Subtag AssemblyRequirements: Subtags of this tag define the set of requirements that will be used when evaluating SEWING assemblies

Subtag ClashRequirement: Checks for clashes between segments in the assembly

maximum_clashes_allowed: Maximum number of clashes to allow in the assembly
clash_radius: Radius in Angstroms within which two residues are considered to be clashing

Subtag DsspSpecificLengthRequirement: Restricts the number of residues in segments with the specified DSSP

dssp_code: DSSP code whose length the requirement is restricting
maximum_length: Maximum number of residues in a segment with the given secondary structure
minimum_length: Minimum number of residues in a segment with the given secondary structure

Subtag KeepLigandContactsRequirement: Fails if an assembly's ligands lose more than a set number of contacts

contact_distance_cutoff: Maximum distance between two contact atoms before the contact is considered broken

Subtag LengthInResiduesRequirement: Checks the number of segments in the assembly

maximum_length: Maximum number of residues to allow in the assembly
minimum_length: Minimum number of residues in the final assembly

Subtag LigandClashRequirement: Checks for clashes between the assembly and its ligands

maximum_clashes_allowed: Maximum number of clashes to allow in the assembly
clash_radius: Radius in Angstroms within which two residues are considered to be clashing

Subtag SizeInSegmentsRequirement: Checks the number of segments in the assembly

maximum_size: Maximum number of secondary structure elements (including loops) to allow in the assembly
minimum_size: Minimum number of secondary structure elements (including loops) in the final assembly

Subtag Ligands: Subtags of this tag specify the ligands present in the input pose and their respective protein contacts.

Subtag Ligand: Specifies the position of a ligand and the contacts that it forms with the input pose

partner_ligand: Is this ligand found in the partner PDB?
pdb_conformers: Name of file containing a list of PDBs (or other Rosetta-compatible input files) containing alternate ligand conformations to sample
alignment_atoms: Comma-separated list of atom names to use when aligning ligand conformers to one another
auto_detect_contacts: Should we automatically detect contacts that are joined to the ligand by inter-residue chemical bonds?
ligand_resnum: Residue number of ligand in either PDB or Rosetta numbering
ligand_selector: Residue selector indicating ligand(s) covered in this tag. The name of a previously declared residue selector or a logical expression of AND, NOT (!), OR, parentheses, and the names of previously declared residue selectors. Any capitalization of AND, NOT, and OR is accepted. An exclamation mark can be used instead of NOT. Boolean operators have their traditional priorities: NOT then AND then OR. For example, if selectors s1, s2, and s3 have been declared, you could write: 's1 or s2 and not s3' which would select a particular residue if that residue were selected by s1 or if it were selected by s2 but not by s3.

Subtag Contact:

partner_contact: Does this tag specify a contact with the partner PDB?
contact_resnum: (REQUIRED) Number of residue participating in this contact in PDB or Rosetta numbering
ligand_atom_name: Rosetta name for the ligand atom participating in the contact
contact_atom_name: Rosetta name for the protein atom participating in the contact

Subtag Coordination: Contains subtags defining ideal coordination environments for atoms in the ligand

coordination_files: Comma-separated list of coordination file names for this ligand
geometry_score_threshold: Maximum score geometry score to allow when forming a contact

Subtag IdealContacts:

distance: (REQUIRED) Ideal distance between ligand and contact atom
angle: Ideal angle between this atom's contacts
dihedral_1: Ideal dihedral angle: contact_base - contact - ligand_atom - other_contact
dihedral_2: Ideal dihedral angle: contact - ligand_atom - other_contact - other_base
max_coordinating_atoms: (REQUIRED) Maximum number of contacts that this atom can form. Note that IdealContacts tags do not need to be defined for atoms with no contacts.
ligand_atom_name: (REQUIRED) Rosetta name of the ligand atom to which this tag applies

Binding Partners

AppendAssemblyMover allows users to specify an additional PDB as a binding partner for their starting structure using the partner_pdb attribute of the AppendAssemblyMover tag. The structure provided in this PDB file must already have the correct coordinates relative to the starting structure provided to AppendAssemblyMover (no docking or minimization occurs before or during the protocol). The ClashRequirement will automatically check for clashes between the assembly and the binding partner.

In most cases, users will also want to include the PartnerMotifScorer in their set of AssemblyScorers. This scorer evaluates possible packing between the assembly and the binding partner. In addition to the standard recommended scorers and requirements, other potentially useful scorers and requirements for dealing with partner PDBs include:

SubsetPartnerMotifScorer: Evaluates possible packing between the assembly and a specified region of the partner PDB

The output_partner attribute of AppendAssemblyMover (default true) toggles whether the binding partner will be included in PDB files output from AppendAssemblyMover. Note that the binding partner is not modified or moved during assembly.

Ligands

The Ligands subtag is a new feature of AppendAssemblyMover which allows users to designate non-protein residues in their starting structure as ligand molecules. AppendAssemblyMover stores and tracks which residues in the starting node are in contact with the ligand and automatically designates them as required residues.

If ligands are included in an assembly, it is highly recommended that users include the LigandClashRequirement as the ClashRequirement does not detect clashes with ligands. Users may also want to include the LigandScorer in their AssemblyScorers section; this scorer evaluates hydrophobic packing between the assembly and the ligand.

The syntax for defining a ligand is as follows:

<Ligands>
  <Ligand ligand_resnum="(&core::Size;)" ligand_selector="(&string;)" auto_detect_contacts="(true &bool;) pdb_conformers="(&string;) alignment_atoms="(&string;)>
    <Contact contact_resnum="(&core::Size;)" contact_atom_name="(&string;)" ligand_atom_name="(&string;)" />
    . . . (additional Contact subtags)
  </Ligand>
 . . . (additional Ligand subtags)
</Ligands>

Each AppendAssemblyMover may have a Ligands subtag which can contain one or more Ligand subtags. Each ligand can be specified either using a residue number or using a residue selector.

The auto_detect_contacts option (default true) indicates that AppendAssemblyMover should automatically detect contacts between the ligand and the starting structure. Currently, only covalent bonds and protein-metal contacts (including contacts to metal atoms in larger ligands) can be detected automatically. We intend to add support for hydrogen bond detection in the future. If auto_detect_contacts is false or if no detectable contacts exist, the user MUST manually specify at least one contact between the ligand and the starting structure.

Ligand contacts may also be manually specified using Contact subtags. Each Contact tag must specify the names of the two atoms involved in the contact as well as the residue number within the starting structure (Rosetta or PDB numbering) of the contact residue.

AppendAssemblyMover also includes support for ligand conformer sampling. In order to enable conformer sampling, three steps must be followed:

1) The user must set the conformer_switch_probability attribute of the AppendAssemblyMover tag to a nonzero value (conformer sampling is turned off by default). 2) The user must provide a text file containing a list of PDB conformers using the pdb_conformers attribute of the Ligand tag. Each PDB file should contain one ligand conformation with no additional structure/residues. 3) The user must specify three ligand atom names using the alignment_atoms attribute of the Ligand tag. These atoms will be used to superimpose conformers onto one another during conformer switching. It is highly recommended that the user select atoms that will retain the geometry of any atoms involved in contacts.

If conformer sampling is enabled, it is highly recommended that the user include the KeepLigandContactsRequirement in the set of requirements. This requirement ensures that conformer sampling does not increase the distance between two contact atoms beyond a certain user-defined threshold.

Example

The following is an example RosettaScript using AppendAssemblyMover:

<ROSETTASCRIPTS>
  <SCOREFXNS>
  </SCOREFXNS>
  <RESIDUE_SELECTORS>
    <ResidueName name="select_zn" residue_name3=" ZN" />
  </RESIDUE_SELECTORS>
  <FILTERS>
  </FILTERS>
  <MOVERS>
    <AppendAssemblyMover name="assemble" model_file_name="/nas02/home/g/u/guffy/smotifs_H_5_40_L_1_6_H_5_40.segments" add_probability="0.05" hashed="false" minimum_cycles="10000" maximum_cycles="20000" start_temperature="1.5" end_temperature="0.1">
      <Ligands>
        <Ligand ligand_selector="select_zn" auto_detect_contacts="true" />
      </Ligands>
      <AssemblyRequirements>
        <DsspSpecificLengthRequirement dssp_code="L" maximum_length="6" /> Prevents super-long loops, should be unnecessary with this segment file                                                                                                       
        <DsspSpecificLengthRequirement dssp_code="H" maximum_length="25" minimum_length="10" /> Prevents super-short helices                                                                                                                             
        <ClashRequirement />
        <LigandClashRequirement />
        <SizeInSegmentsRequirement maximum_size="9" minimum_size="5" />
      </AssemblyRequirements>
    </AppendAssemblyMover>
  </MOVERS>
  <APPLY_TO_POSE>
  </APPLY_TO_POSE>
  <PROTOCOLS>
        <Add mover_name="assemble" />
  </PROTOCOLS>
</ROSETTASCRIPTS>