The Peptide in Protein Annotation node annotates values that depend on the position of the identified peptides and the proteins found in a sample. The node can optionally add the Annotated Sequence column, the Modifications in Master Protein column, and the Modifications in All Proteins column to the Peptide Groups page and the Proteins page in the result file to display these annotations. The node also automatically adds the Sequence Coverage column to the Proteins page.

Specifically, the Peptide in Protein Annotation node annotates the following values:

  • The position of the identified peptide in the proteins containing the peptide.
  • The flanking residues of the peptide.
  • The positions of the found modifications in the protein sequences where the peptide is located.
  • The concatenated list of all modification positions found for the protein.

All these values depend on the position of the identified peptides in the proteins.

For information on these values and the Peptide in Protein Annotation workflow, see Adding PTM Analysis to a Workflow.

The flanking residues of a peptide are the amino acids that border a cleavage site. The Annotate Flanking Residues of the Peptide parameter of the Peptide in Protein Annotation node determines whether the Proteome Discoverer application annotates the amino acids before and after the peptide in the protein sequence. When you set this parameter to True, the application adds the Annotated Sequence column to the Peptide Groups page. The column displays the one-letter code representing the flanking residues of a peptide in a protein.

The following table describes the parameters for the Peptide in Protein Annotation node.

Peptide in Protein Annotation node parameters

Parameter

Definition

Annotate Flanking Residues of the Peptide

Determines whether to annotate the amino acids before and after the peptide in the protein sequence.

  • (Default) True: Annotates the amino acids before and after the peptide in the protein sequence. It adds the Annotated Sequence column containing this annotation information to the Peptide Groups page of the result file. The application separates the names of the amino acids before and after the peptide sequence with a dot. There might be more than one letter if the peptide has different flanking residues in different proteins.
  • False: Does not annotate the amino acids before and after the peptide, and does not add the Annotated Sequence column to the Peptide Groups page of the result file.

Number Flanking Residues in Connection Tables

Specifies the number of flanking residues to report in the associated tables opened by clicking Show Associated Tables in the result file. The number of flanking residues in the primary Peptide Groups table is always 1.

Range: 1–5; default: 1

Protein Modifications Reported

Determines which protein modifications are displayed in the Position in Master Proteins column in the Peptide Groups page of the result file.

  • None: Does not annotate the positions for any proteins.
  • (Default) Only for Master Proteins: Annotates only the positions of the master protein of each protein group.
  • For All Proteins: Annotates the positions of all proteins.

Modification Sites Reported

Determines which modifications to annotate on the Proteins page of the result file.

  • None: Does not annotate any protein modification sites.
  • (Default) All Combined: Annotates all modification sites and displays the annotations in a single column called Modifications. Peptide terminal modifications are not transferred to the protein level.
  • Only Specific: Annotates only the modification sites specified by the parameters of this node and displays this information in two columns, Modification_name and Modification_name Count. The first column shows the position of all identified modifications of this kind in the protein. The second column contains the total count of this kind of modification in each protein.
  • All and Specific: Annotates all identified modifications of the protein and displays the annotations in a single column called Modifications. It also annotates selected modifications and displays the annotations in separate single columns.

Minimum PSM Confidence

Specifies what to consider as the lowest PSM confidence level of the PSMs connected to a protein:

  • Low: Considers low confidence the lowest PSM confidence.
  • Medium: Considers medium confidence the lowest PSM confidence.
  • (Default) High: Considers high confidence the lowest PSM confidence.

Report Only PTMs

Determines how to display peptide modifications on the Peptide Groups page.

  • (Default) True: Displays only the modifications that are classified as PTMs for the target amino acid that they were identified for. The classifications for the amino acid/modification pairs are defined in the view opened by the Administration > Maintain Chemical Modifications command.
  • The following classifications are treated as PTMs:
    - Post-translational
    - Pretranslational
    - N-linked glycosylation
    - O-linked glycosylation
    - Other glycosylation
    - Multiple
  • False: Displays all modifications.

N-Terminal Modification

Selects any known or suspected N-terminal modification.

Default: None

C-Terminal Modification

Selects any known or suspected C-terminal modification.

Default: None

Modification

Selects any known or suspected modifications.

Default: None

Protein Positions for Peptides

Determines which protein modification positions the application annotates on the Peptide Groups page of the result.

  • None: Does not annotate the positions for any proteins.
  • (Default) Only for Master Proteins: Annotates only the positions of the master protein of each protein group and displays the annotations in the Positions in Master Proteins column to the Peptide Groups page.
  • It displays these positions in the following form:
  • protein accession [start - end]
  • For All Proteins: Annotates the positions of all proteins and displays the annotations in the Position in All Proteins column to the Peptide Groups page.