SSRCalc

SSRCalc predicts retention times of tryptic peptides in reversed-phase high-performance liquid chromatography (HPLC) coupled with matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) to support peptide mapping and discrimination of peptides with similar mass-to-charge ratios.

Key Features:

• Sequence-specific model: Calculates peptide retention time from sequence information by summing amino-acid-specific retention coefficients.
• N-terminal coefficients: Includes additional terms that account for retention coefficients of amino acids at the peptide N-terminus.
• Coefficient sets: Utilizes two adjustable sets of coefficients optimized during model fitting.
• Length and hydrophobicity compensation: Incorporates compensations for peptide length and overall hydrophobicity.
• Training data: Model optimization used 346 tryptic peptides in the 560–4,000 Da mass range derived from 17 protein digests.
• Chromatography conditions: Applicable to reversed-phase columns with 300-Å pore size using linear water–acetonitrile (ACN) gradients and trifluoroacetic acid (TFA) as the ion-pairing modifier.
• Injection and column formats: Validated for direct sample injection and pre-column injection and across nano- to narrow-bore column sizes.
• Quantitative performance: Produces a robust linear relationship between retention time and hydrophobicity with R² ≈ 0.94.

Scientific Applications:

• Distinguishing close m/z peptides: Aids discrimination of peptides with close mass-to-charge (m/z) values by providing predicted retention times.
• Peptide mapping: Supports detailed peptide mapping of selected proteins in proteomics workflows.
• HPLC-MALDI-MS analyses: Enhances retention time prediction accuracy for HPLC coupled to MALDI MS across column sizes and injection modes.

Methodology:

The method sums amino-acid-specific retention coefficients with additional N-terminal terms, fits two sets of coefficients while applying compensations for peptide length and hydrophobicity, and was optimized on 346 tryptic peptides (560–4,000 Da) from 17 protein digests yielding a linear relationship with R² ≈ 0.94.