public static void OGlycoTest_FragmentIons2()
{
//Get glycanBox
var glycanBox = OGlycanBoxes[24];
Protein protein = new Protein("TVYLGASK", "");
var peptide = protein.Digest(new DigestionParams(), new List <Modification>(), new List <Modification>()).First();
List <int> modPos = new List <int> {
2, 8
};
var peptideWithMod = GlycoPeptides.OGlyGetTheoreticalPeptide(modPos.ToArray(), peptide, OGlycanBoxes[24]);
Assert.That(peptideWithMod.FullSequence == "T[O-Glycosylation:H1N1 on X]VYLGAS[O-Glycosylation:H1N1A1 on X]K");
var fragments_etd = GlycoPeptides.OGlyGetTheoreticalFragments(DissociationType.ETD, peptide, peptideWithMod);
Assert.That(fragments_etd.Count == 22);
Assert.That(fragments_etd.Last().Annotation == "zDot8");
Assert.That(fragments_etd.Last().NeutralMass > 1824);
var fragments_hcd = GlycoPeptides.OGlyGetTheoreticalFragments(DissociationType.HCD, peptide, peptideWithMod);
Assert.That(fragments_hcd.Where(p => p.ProductType == ProductType.M).Count() == 5);
var fragments_ethcd = GlycoPeptides.OGlyGetTheoreticalFragments(DissociationType.EThcD, peptide, peptideWithMod);
Assert.That(fragments_ethcd.Where(p => p.ProductType == ProductType.M).Count() == 5);
}
public static void OxoniumIonAnalysis()
{
Assert.That(Glycan.AllOxoniumIons[4] == 13805550);
Assert.That(Glycan.AllOxoniumIons[5] == 14406607);
Assert.That(Glycan.AllOxoniumIons[9] == 20408720);
Assert.That(Glycan.AllOxoniumIons[10] == 27409268);
Assert.That(Glycan.AllOxoniumIons[12] == 29210324);
Assert.That(Glycan.AllOxoniumIons[14] == 36614002);
//Get Scan
CommonParameters commonParameters = new CommonParameters(dissociationType: DissociationType.EThcD, trimMsMsPeaks: false);
string spectraFile = Path.Combine(TestContext.CurrentContext.TestDirectory, @"GlycoTestData\2019_09_16_StcEmix_35trig_EThcD25_rep1_4565.mgf");
var file = new MyFileManager(true).LoadFile(spectraFile, commonParameters);
var scan = MetaMorpheusTask.GetMs2Scans(file, spectraFile, commonParameters).First();
var productSearchMode = new SinglePpmAroundZeroSearchMode(20);
var oxoniumIonIntensities = GlycoPeptides.ScanOxoniumIonFilter(scan, productSearchMode, DissociationType.EThcD);
//Get glycanBox
var glycanBox = OGlycanBoxes[19];
var satifyOxonium = GlycoPeptides.OxoniumIonsAnalysis(oxoniumIonIntensities, glycanBox);
Assert.That(satifyOxonium);
}
public static void GlyTest_GlyGetTheoreticalFragments()
{
Protein pep = new Protein("TKPREEQYNSTYR", "accession");
DigestionParams digestionParams = new DigestionParams(minPeptideLength: 7);
var aPeptideWithSetModifications = pep.Digest(digestionParams, new List <Modification>(), new List <Modification>());
string[] motifs = new string[] { "Nxs", "Nxt" };
var sites = GlycoSpectralMatch.GetPossibleModSites(aPeptideWithSetModifications.Last(), motifs);
Glycan glycan = Glycan.Struct2Glycan("(N(F)(N(H(H(N))(H(N)))))", 0);
//using (StreamWriter output = new StreamWriter(Path.Combine(TestContext.CurrentContext.TestDirectory, "GlycanFragmentions.txt")))
//{
// foreach (var product in fragmentIons)
// {
// foreach (var ion in product.Item2)
// {
// output.WriteLine(ion.Annotation + "\t" + ion.NeutralLoss.ToString() + "\t" + ion.NeutralMass.ToString());
// }
// }
//}
CommonParameters commonParameters = new CommonParameters(deconvolutionMassTolerance: new PpmTolerance(20), trimMsMsPeaks: false);
string filePath = Path.Combine(TestContext.CurrentContext.TestDirectory, @"GlycoTestData/Glyco_3383.mgf"); //"25170.mgf"
MyFileManager myFileManager = new MyFileManager(true);
var msDataFile = myFileManager.LoadFile(filePath, commonParameters);
var listOfSortedms2Scans = MetaMorpheusTask.GetMs2Scans(msDataFile, filePath, commonParameters).ToArray();
var glycanMod = Glycan.NGlycanToModification(glycan);
var glycopep = GlycoPeptides.GenerateGlycopeptide(sites[0], aPeptideWithSetModifications.Last(), glycan);
List <Product> fragmentIons = new List <Product>();
glycopep.Fragment(DissociationType.HCD, FragmentationTerminus.Both, fragmentIons);
var glycanYIons = GlycoPeptides.GetGlycanYIons(listOfSortedms2Scans[0].PrecursorMass, glycan);
var matchedGlycanYIons = MetaMorpheusEngine.MatchFragmentIons(listOfSortedms2Scans[0], glycanYIons, commonParameters);
Assert.AreEqual(matchedGlycanYIons.Count, 14);
//TO DO: The neutroloss is not annotated well.
var matchedFragmentIons = MetaMorpheusEngine.MatchFragmentIons(listOfSortedms2Scans[0], fragmentIons, commonParameters);
var coreIons = GlycoPeptides.ScanGetTrimannosylCore(matchedFragmentIons, glycan);
Assert.AreEqual(coreIons.Count, 6);
var filter = GlycoPeptides.ScanTrimannosylCoreFilter(matchedFragmentIons, glycan);
Assert.AreEqual(filter, true);
var NGlycans = GlycanDatabase.LoadGlycan(GlobalVariables.NGlycanLocations[0], true, false);
var bestGlycans = GlycoPeptides.MatchBestGlycan(listOfSortedms2Scans[0], NGlycans.ToArray(), commonParameters).Where(p => p != null && p.Item2 >= 2).OrderByDescending(p => p.Item2).Take(100).OrderBy(p => p.Item3).ToArray();;
}
public static void OGlycoTest_GetKPerWithDuplicate()
{
List <int> input = new List <int> {
3, 5, 2, 7
};
int[] ids = new int[3] {
2, 2, 3
};
var perWithDuplicate = GlycoPeptides.GetPermutations(input, ids);
var allPermutation = Glycan.GetPermutations(input, ids.Length);
Assert.That(perWithDuplicate.Count() == allPermutation.Count() / 2);
}
public static void OGlycoTest_FragmentIonsHash()
{
//Get glycanBox
var glycanBox = OGlycanBoxes[8];
Protein protein = new Protein("PTLFKNVSLYK", "");
var peptide = protein.Digest(new DigestionParams(), new List <Modification>(), new List <Modification>()).First();
List <int> modPos = GlycoSpectralMatch.GetPossibleModSites(peptide, new string[] { "S", "T" });
var peptideWithMod = GlycoPeptides.OGlyGetTheoreticalPeptide(modPos.ToArray(), peptide, glycanBox);
Assert.That(peptideWithMod.FullSequence == "PT[O-Glycosylation:N1A1 on X]LFKNVS[O-Glycosylation:N1 on X]LYK");
//The following code prove that the default Fragment method doesn't work for O-glycopeptide due to neutral losses.
var fragments_hcd = new List <Product>();
peptide.Fragment(DissociationType.HCD, FragmentationTerminus.Both, fragments_hcd);
var fragmentsMod_hcd = new List <Product>();
peptideWithMod.Fragment(DissociationType.HCD, FragmentationTerminus.Both, fragmentsMod_hcd);
Assert.That(fragments_hcd.Count() == 20);
Assert.That(fragmentsMod_hcd.Count() == 61); //The Fragments also contain neutral loss ions.
var frag_ments_etd = new List <Product>();
peptide.Fragment(DissociationType.ETD, FragmentationTerminus.Both, frag_ments_etd);
var fragmentsMod_etd = new List <Product>();
peptideWithMod.Fragment(DissociationType.ETD, FragmentationTerminus.Both, fragmentsMod_etd);
//Tuple<int, int[]> keyValuePair represents: <glycanBoxId, glycanModPositions>
Tuple <int, int[]> keyValuePairs = new Tuple <int, int[]>(8, modPos.ToArray());
var fragments_etd_origin = GlycoPeptides.GetFragmentHash(frag_ments_etd, new Tuple <int, int[]>(0, null), OGlycanBoxes, 1000);
var fragmentsHash_etd = GlycoPeptides.GetFragmentHash(frag_ments_etd, keyValuePairs, OGlycanBoxes, 1000);
var fragmentsMod_etd_origin = GlycoPeptides.GetFragmentHash(fragmentsMod_etd, new Tuple <int, int[]>(0, null), OGlycanBoxes, 1000);
var overlap = fragmentsHash_etd.Intersect(fragments_etd_origin).Count();
Assert.That(overlap == 14);
var overlap2 = fragmentsHash_etd.Intersect(fragmentsMod_etd_origin).Count();
//ETD didn't change y ions.
Assert.That(overlap2 == 23);
}
public static void OGlycoTest_FragmentIons()
{
//This test is to test ETD on proline. As proline didn't have proline related product ions.
//Get glycanBox
var glycanBox = OGlycanBoxes[8];
Protein protein = new Protein("PTLFKNVSLYK", "");
var peptide = protein.Digest(new DigestionParams(), new List <Modification>(), new List <Modification>()).First();
List <int> modPos = GlycoSpectralMatch.GetPossibleModSites(peptide, new string[] { "S", "T" });
var peptideWithMod = GlycoPeptides.OGlyGetTheoreticalPeptide(modPos.ToArray(), peptide, OGlycanBoxes[8]);
Assert.That(peptideWithMod.FullSequence == "PT[O-Glycosylation:N1A1 on X]LFKNVS[O-Glycosylation:N1 on X]LYK");
var fragments_hcd = GlycoPeptides.OGlyGetTheoreticalFragments(DissociationType.HCD, peptide, peptideWithMod);
var fragments_ethcd = GlycoPeptides.OGlyGetTheoreticalFragments(DissociationType.EThcD, peptide, peptideWithMod);
}
public static void OGlycoTest_Localization2()
{
//There may have a bug that MM cannot identify Peptide modified with (HexNAc), This is to test and find the bug.
//Get glycanBox
var glycanBox = OGlycanBoxes[0];
//Get unmodified peptide, products, allPossible modPos and all boxes.
Protein protein = new Protein("AATVGSLAGQPLQER", "P16150");
var peptide = protein.Digest(new DigestionParams(), new List <Modification>(), new List <Modification>()).First();
List <Product> products = new List <Product>();
peptide.Fragment(DissociationType.ETD, FragmentationTerminus.Both, products);
int[] modPos = GlycoSpectralMatch.GetPossibleModSites(peptide, new string[] { "S", "T" }).OrderBy(p => p).ToArray();
var boxes = GlycanBox.BuildChildOGlycanBoxes(glycanBox.NumberOfMods, glycanBox.ModIds).ToArray();
//Load scan.
CommonParameters commonParameters = new CommonParameters(dissociationType: DissociationType.ETD, trimMsMsPeaks: false);
string spectraFile = Path.Combine(TestContext.CurrentContext.TestDirectory, @"GlycoTestData\181217_Fusion_(LC2)_NewObj_Serum_deSA_Jacalin_HRM_4h_ETD_HCD_DDA_mz(400_1200)_21707.mgf");
var file = new MyFileManager(true).LoadFile(spectraFile, commonParameters);
var scans = MetaMorpheusTask.GetMs2Scans(file, spectraFile, commonParameters).ToArray();
//Known peptideWithMod match.
var peptideWithMod = GlycoPeptides.OGlyGetTheoreticalPeptide(new int[1] {
4
}, peptide, glycanBox);
Assert.That(peptideWithMod.FullSequence == "AAT[O-Glycosylation:N1 on X]VGSLAGQPLQER");
//List<Product> knownProducts = peptideWithMod.Fragment(DissociationType.EThcD, FragmentationTerminus.Both).ToList();
List <Product> knownProducts = GlycoPeptides.OGlyGetTheoreticalFragments(DissociationType.ETD, peptide, peptideWithMod);
var matchedKnownFragmentIons = MetaMorpheusEngine.MatchFragmentIons(scans.First(), knownProducts, commonParameters);
//Get hashset int
int obsPreviousFragmentCeilingMz = 0;
List <int> binsToSearch = new List <int>();
foreach (var envelope in scans.First().ExperimentalFragments)
{
// assume charge state 1 to calculate mass tolerance
double experimentalFragmentMass = envelope.MonoisotopicMass;
// get theoretical fragment bins within mass tolerance
int obsFragmentFloorMass = (int)Math.Floor((commonParameters.ProductMassTolerance.GetMinimumValue(experimentalFragmentMass)) * 1000);
int obsFragmentCeilingMass = (int)Math.Ceiling((commonParameters.ProductMassTolerance.GetMaximumValue(experimentalFragmentMass)) * 1000);
// prevents double-counting peaks close in m/z and lower-bound out of range exceptions
if (obsFragmentFloorMass < obsPreviousFragmentCeilingMz)
{
obsFragmentFloorMass = obsPreviousFragmentCeilingMz;
}
obsPreviousFragmentCeilingMz = obsFragmentCeilingMass + 1;
// search mass bins within a tolerance
for (int fragmentBin = obsFragmentFloorMass; fragmentBin <= obsFragmentCeilingMass; fragmentBin++)
{
binsToSearch.Add(fragmentBin);
}
}
HashSet <int> allPeaks = new HashSet <int>(binsToSearch);
//Graph Localization
LocalizationGraph localizationGraph = new LocalizationGraph(modPos, glycanBox, boxes, -1);
LocalizationGraph.LocalizeOGlycan(localizationGraph, scans.First(), commonParameters.ProductMassTolerance, products);
var allPaths = LocalizationGraph.GetAllHighestScorePaths(localizationGraph.array, localizationGraph.ChildModBoxes);
var knowPath = new int[2] {
1, 1
};
Assert.That(Enumerable.SequenceEqual(knowPath, allPaths[0]));
var local = LocalizationGraph.GetLocalizedPath(localizationGraph, allPaths.First());
Assert.That(Enumerable.SequenceEqual(local.Mods.Select(p => p.Item1), new List <int> {
4
}));
Assert.That(Enumerable.SequenceEqual(local.Mods.Select(p => p.Item2), new List <int> {
0
}));
}
public static void OGlycoTest_Localization()
{
//Get glycanBox
var glycanBox = OGlycanBoxes[19];
//Get unmodified peptide, products, allPossible modPos and all boxes.
Protein protein = new Protein("TTGSLEPSSGASGPQVSSVK", "P16150");
var peptide = protein.Digest(new DigestionParams(), new List <Modification>(), new List <Modification>()).First();
List <Product> products = new List <Product>();
peptide.Fragment(DissociationType.ETD, FragmentationTerminus.Both, products);
int[] modPos = GlycoSpectralMatch.GetPossibleModSites(peptide, new string[] { "S", "T" }).OrderBy(v => v).ToArray();
var boxes = GlycanBox.BuildChildOGlycanBoxes(3, glycanBox.ModIds).ToArray();
Assert.That(boxes.Count() == 6);
//Get Unlocal Fragment
var unlocalCost = GlycoPeptides.GetUnlocalFragment(products, modPos, glycanBox);
Assert.That(unlocalCost.Count == 4); //Basicly, the unlocal are c/z ions that don't localize glycosylation.
//Get scan
CommonParameters commonParameters = new CommonParameters(dissociationType: DissociationType.EThcD, trimMsMsPeaks: false);
string spectraFile = Path.Combine(TestContext.CurrentContext.TestDirectory, @"GlycoTestData\2019_09_16_StcEmix_35trig_EThcD25_rep1_4565.mgf");
var file = new MyFileManager(true).LoadFile(spectraFile, commonParameters);
var scans = MetaMorpheusTask.GetMs2Scans(file, spectraFile, commonParameters).ToArray();
//Known peptideWithMod match.
var peptideWithMod = GlycoPeptides.OGlyGetTheoreticalPeptide(new int[3] {
10, 2, 3
}, peptide, glycanBox);
Assert.That(peptideWithMod.FullSequence == "T[O-Glycosylation:H1N1 on X]T[O-Glycosylation:H1N1 on X]GSLEPSS[O-Glycosylation:N1 on X]GASGPQVSSVK");
List <Product> knownProducts = GlycoPeptides.OGlyGetTheoreticalFragments(DissociationType.EThcD, peptide, peptideWithMod);
var matchedKnownFragmentIons = MetaMorpheusEngine.MatchFragmentIons(scans.First(), knownProducts, commonParameters);
//Graph Localization
LocalizationGraph localizationGraph = new LocalizationGraph(modPos, glycanBox, boxes, -1);
LocalizationGraph.LocalizeOGlycan(localizationGraph, scans.First(), commonParameters.ProductMassTolerance, products);
var allPaths = LocalizationGraph.GetAllHighestScorePaths(localizationGraph.array, localizationGraph.ChildModBoxes);
var knowPath = new int[8] {
2, 4, 4, 4, 5, 5, 5, 5
};
Assert.That(Enumerable.SequenceEqual(knowPath, allPaths[0]));
//Get localized Route
var local = LocalizationGraph.GetLocalizedPath(localizationGraph, allPaths.First());
Assert.That(Enumerable.SequenceEqual(local.Mods.Select(v => v.Item1), new List <int> {
2, 3, 10
}));
Assert.That(Enumerable.SequenceEqual(local.Mods.Select(v => v.Item2), new List <int> {
1, 1, 0
}));
//Get all paths, calculate PScore and calculate position probability.
var p = scans.First().TheScan.MassSpectrum.Size *commonParameters.ProductMassTolerance.GetRange(1000).Width / scans.First().TheScan.MassSpectrum.Range.Width;
var n = knownProducts.Where(v => v.ProductType == ProductType.c || v.ProductType == ProductType.zDot).Count();
var allPathWithWeights = LocalizationGraph.GetAllPaths_CalP(localizationGraph, p, n);
Assert.That(allPathWithWeights.Count == 168);
//Calculate Site Specific Localization Probability
var y = LocalizationGraph.CalSiteSpecificLocalizationProbability(allPathWithWeights, localizationGraph.ModPos);
Assert.That(y.Count == 8);
Assert.That(y.First().Value[1].Item2 > 0.99);
}
