In the intact mass analysis workflow, the BioPharma Finder application can match the measured masses of detected components to the masses of user-specified target sequences, aiding in the identification of components. You can set the parameters for target sequence matching in your intact mass analysis experiment on the Parameters > Identification page of the processing method editing wizard. See Edit the Identification page and Use real-time optimization (RTO) to reprocess results.

You can use target sequence matching for intact mass analysis for experiments incorporating:

  • Any deconvolution algorithm: Xtract or ReSpect
  • Any deconvolution method: Average over selected retention time, automatic peak detection, or sliding windows
  • Any results format: Batch Processing or Multiconsensus

The target sequences you add to your experiment can include the following modifications, applied in the listed order:

  1. Disulfide links, which form bonds between cysteine residues in a heavy chain and light chain or within the same chain.
  2. Site-specific static modifications, which are side-chain or terminal modifications that you can apply to a single site for a specified residue, nucleotide base, or terminal.
  3. Global static modifications, which are side-chain or terminal modifications that you can apply to all sites for a specified residue, nucleotide base, or terminal.
  4. Glycosylations, which refers to the attachment of sugar moieties at sites on the target sequence to form glycans.
  5. The application applies all possible combinations between zero and a maximum number of user-specified variable modifications to the target sequence.

  6. NOTE

    The algorithm that detects possible glycosylation sites was designed for use on intact proteins. If it is applied to digested peptides, it might fail to identify motifs that have been truncated by a cleavage.

    To address this issue when using a peptide as a target sequence, append an amino acid to the sequence to complete the motif, and then define and apply a custom modification that subtracts the mass of that amino acid.

  7. Variable modifications, which are possible side-chain or terminal modifications whose specific sites and number of occurrences might not be known.
  8. The application applies all possible combinations between zero and a maximum number of user-specified variable modifications to the target sequence.
  9. When you select glycosylations and variable modifications, the application also generates additional potential component matches in cases where glycosylations and variable modifications occur together.
  10. See Assigning modifications to a protein sequence.