Expert sytem for annotation of MS/MS spectra - Supplementary Table 1 rule name condition if-action else-action. 1 ppm or Da if mass analyzer was Orbitrap.
ppm or Da Ready for immonium ions Ready for side chain ions immonium Ala immonium Arg immonium Asn immonium Asp immonium Cys immonium CamC immonium Gln immonium Glu immonium Gly immonium His immonium Ile immonium Leu immonium Lys immonium Met immonium Met(Ox) immonium Phe immonium Pro immonium Ser immonium phSer immonium Thr immonium phThr immonium Trp immonium Tyr immonium phTyr immonium Val side chain by R side chain by H side chain by K side chain by W Parent ion a-ion series b-ion series y-ion series stable c1 stable x by Phosphorylation General Enumeration start remove unstable ions unstable a1 unstable a2 unstable b1 Charge 1+ Charge 2+ Charge 3+ Charge 4+ Charge 5+ Neutral loss at phS Neutral loss at phT Neutral loss at phY Ready for Neutral loss Ready for Internal Fragmentation internal-fragment (P) internal-fragment (H) internal-fragment (E) internal-fragment (D) internal-fragment (F) internal-fragment (S) internal-fragment (L) internal-fragment (K) internal-fragment (A) internal-fragment (R) internal-fragment (N) internal-fragment (C) internal-fragment (Q) internal-fragment (G) internal-fragment (I) internal-fragment (M) internal-fragment (T) internal-fragment (W) internal-fragment (Y) internal-fragment (V) Neutral loss at C Neutral loss at D Neutral loss at E Neutral loss at I Neutral loss at K Neutral loss at L Neutral loss at M Neutral loss at M(Ox) Neutral loss at N Neutral loss at Q Neutral loss at R Neutral loss at S
86 87 88 89 90
Neutral loss at T Neutral loss at V Neutral loss at W Neutral loss at IM K-NH3 Neutral loss at C-Term-H2O
condition
if-action
else-action
if mass analyzer was Orbitrap if mass analyzer was Orbitrap if mass analyzer was Orbitrap if rule2 is true and peptide sequence contains A and m/z value in spectrum if rule2 is true and peptide sequence contains R and m/z value in spectrum if rule2 is true and peptide sequence contains N and m/z value in spectrum if rule2 is true and peptide sequence contains D acid and m/z value in spectrum if rule2 is true and peptide sequence contains C without modification on Cysteine and m/z value in spectrum if rule2 is true and peptide sequence contains C with modification of Carbamidometyl and m/z value in spectrum if rule2 is true and peptide sequence contains E and m/z value in spectrum if rule2 is true and peptide sequence contains Q and m/z value in spectrum if rule2 is true and peptide sequence contains G and m/z value in spectrum if rule2 is true and peptide sequence contains H and m/z value in spectrum if rule2 is true and peptide sequence contains I and m/z value in spectrum if rule2 is true and peptide sequence contains L and m/z value in spectrum if rule2 is true and peptide sequence contains K and m/z value in spectrum if rule2 is true and peptide sequence contains M without modification on Methionine and m/z value in spectrum if rule2 is true and peptide sequence contains M with modification of Oxidation and m/z value in spectrum if rule2 is true and peptide sequence contains F and m/z value in spectrum if rule2 is true and peptide sequence contains P and m/z value in spectrum if rule2 is true and peptide sequence contains S and m/z value in spectrum if rule2 is true and peptide sequence contains S with modification of Phosphorylation and m/z value in spectrum if rule2 is true and peptide sequence contains T and m/z value in spectrum if rule2 is true and peptide sequence contains T with modification of Phosphorylation and m/z value in spectrum if rule2 is true and peptide sequence contains W and m/z value in spectrum if rule2 is true and peptide sequence contains Y and m/z value in spectrum if rule2 is true and peptide sequence contains Y with modification of Phosphorylation and m/z value in spectrum if rule2 is true and peptide sequence contains Y and m/z value in spectrum if rule3 is true and peptide sequence contains R and m/z value in spectrum if rule3 is true and peptide sequence contains H and m/z value in spectrum if rule3 is true and peptide sequence contains K and m/z value in spectrum if rule3 is true and peptide sequence contains T and m/z value in spectrum if fragmentation is HCD or CID if fragmentation is HCD or CID if fragmentation is HCD or CID if fragmentation is HCD or CID if second amino acid is a Gln or Asn if peptide is phosphorylated if there is an element in the queue if annotation is declared as unstable if annotation is a1 and has no modification if annotation is a2 and contains Q or N or has a Carbamidometylated C if annotation is b1 and peptide sequence is not starting with R, H, K or Acetylated M,S, A if annotation charge is 1+ and peptide charge is greater than or equals 1+ and m/z value in spectrum if annotation charge is 2+ and peptide charge is greater than or equals 2+ and m/z value in spectrum if annotation charge is 3+ and peptide charge is greater than or equals 3+ and m/z value in spectrum if annotation charge is 4+ and peptide charge is greater than or equals 4+ and m/z value in spectrum if annotation charge is 5+ and peptide charge is greater than or equals 5+ and m/z value in spectrum if fragment sequence contains S and has a Phosphorylation on S and neutral loss count is less than 3 if fragment sequence contains T and has a Phosphorylation on T and neutral loss count is less than 3 if fragment sequence contains Y and has a Phosphorylation on Y and neutral loss count is less than 3 if annotation is in spectrum and neutral loss count is less than 3 if annotation is from a-,b- or y-series if rule50 is true and fragment sequence starts with P if rule50 is true and fragment sequence starts with H if rule50 is true and fragment sequence starts with E if rule50 is true and fragment sequence starts with D if rule50 is true and fragment sequence starts with F if rule50 is true and fragment sequence starts with S if rule50 is true and fragment sequence starts with L if rule50 is true and fragment sequence starts with K if rule50 is true and fragment sequence starts with A if rule50 is true and fragment sequence starts with R if rule50 is true and fragment sequence starts with N if rule50 is true and fragment sequence starts with C if rule50 is true and fragment sequence starts with Q if rule50 is true and fragment sequence starts with G if rule50 is true and fragment sequence starts with I if rule50 is true and fragment sequence starts with M if rule50 is true and fragment sequence starts with T if rule50 is true and fragment sequence starts with W if rule50 is true and fragment sequence starts with Y if rule50 is true and fragment sequence starts with V if rule49 is true and fragment sequence contains C and there is no modification on C if rule49 is true and fragment sequence contains D and there is no modification on D if rule49 is true and fragment sequence contains E and there is no modification on E if rule49 is true and fragment sequence contains I and there is no modification on I if rule49 is true and fragment sequence contains K and there is no modification on K if rule49 is true and fragment sequence contains L and there is no modification on L if rule49 is true and fragment sequence contains M and there is no modification on M if rule49 is true and fragment sequence contains M and M has Oxidation if rule49 is true and fragment sequence contains N and there is no modification on N if rule49 is true and fragment sequence contains Q and there is no modification on Q if rule49 is true and fragment sequence contains R and there is no modification on R if rule49 is true and fragment sequence contains S and there is no modification on S
then fragment tolerance is 20 ppm then immonium ions are possible then side chain loss fragments are possible then add immonium ion of Alanine to candidate list then add immonium ion of Arginine to candidate list then add immonium ion of Asparagine to candidate list then add immonium ion of Aspartic Acid to candidate list then add immonium ion of Cysteine to candidate list then add immonium ion of Carbamidometylated Cysteine to candidate list then add immonium ion of Glutamic acid to candidate list then add immonium ion of Glutamine to candidate list then add immonium ion of Glycine to candidate list then add immonium ion of Histidine to candidate list then add immonium ion of Isoleucine to candidate list then add immonium ion of Leucine to candidate list then add immonium ion of Lysine to candidate list and to queue then add immonium ion of Methionine to candidate list then add immonium ion of Oxidated Methionine to candidate list then add immonium ion of Phenylalanine to candidate list then add immonium ion of Proline to candidate list then add immonium ion of Serine to candidate list then add immonium ion of Phosphorylated Serine to candidate list then add immonium ion of Threonine to candidate list then add immonium ion of Phosphorylated Threonine to candidate list then add immonium ion of Tryptophan to candidate list then add immonium ion of Tyrosine to candidate list then add immonium ion of Phosphorylated Tyrosine to candidate list then add immonium ion of Valine to candidate list then add side chain loss of C4H6N2 to candidate list then add side chain loss of C4H6N2 to candidate list then add side chain loss of C5H11N to candidate list then add side chain losses of C10H9N, C9H9N and C9H7N to candidate list add parent ion to queue add a ion series to queue add b ion series to queue add y ion series to queue add c1 ion to queue add x ions to queue get next elements in the queue discard annotation and remove from queue and go to rule39 a1 is unstable go to rule40 a2 is unstable go to rule40 b1 is unstable go to rule40 add annotation to candidate list increase charge to 2+ and add annotation to candidate list increase charge to 3+ and add annotation to candidate list increase charge to 4+ and add annotation to candidate list increase charge to 5+ and add annotation to candidate list then add annotation with neutral losses of H3PO4 and HPO3 from Phosphorylated S to the queue then add annotation with neutral losses of H3PO4 and HPO3 from Phosphorylated T to the queue then add annotation with neutral losses of H3PO4 and HPO3 from Phosphorylated Y to the queue then annotation can have neutral losses then annotation can have internal fragments then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then generate internal fragments of annotation and add them to the queue then add annotation with neutral loss of CH2S from C to the queue then add annotation with neutral losses of H2O, CO2 and C2H4O2 from D to the queue then add annotation with neutral losses of H2O and C2H4O2 from E to the queue then add annotation with neutral loss of C2H4 from I to the queue then add annotation with neutral losses of C2H5N, C4H9N, C4H11N and C3H9N from K to the queue then add annotation with neutral losses of C3H6 and C4H8 from L to the queue then add annotation with neutral losses of C2H4S and C3H6S from M to the queue then add annotation with neutral losses of CH4SO, C3H8SO and C3H6SO from Oxidated M to the queue then add annotation with neutral losses of NH3, CH3NO and C2H5NO from N to the queue then add annotation with neutral losses of NH3, CH3NO, C2H5NO and C3H5NO from Q to the queue then add annotation with neutral losses of NH3, CH2N2 and C3H9N3 from R to the queue then add annotation with neutral losses of H2O and CH4O from S to the queue
else fragment tolerance is 0.5 Dalton else immonium ions are not possible else side chain loss fragments are not possible
if rule49 is true and fragment sequence contains T and there is no modification on T if rule49 is true and fragment sequence contains V and there is no modification on V if rule49 is true and fragment sequence contains W and there is no modification on W if annotation is immonium ion and fragment sequence contains K and neutral loss count is equals 0 if rule49 is true and (annotation is C-terminal or parent ion) and there is no modification at the C-term and neutral loss count equals 0 91 Neutral loss at N-Term-NH3 if rule49 is true and fragment sequence starts not with P and annotation is N-terminal and there is no modification at the N-term and neutral loss count equals 0 92 Neutral loss at Parent-CO if rule49 is true and annotation is parent ion 93 Neutral loss at internal fragment-CO if rule49 is true and annotation is internal fragment 94 General Enumeration end if there are more elements in the queue 95 Priority Enumeration start if there is an element in the candidate list 96 Filter for null values if annotation is null 97 Priority B Rule if annotation ion type is b 98 Priority Y Rule if annotation ion type is y 99 Priority Parent Rule if annotation ion type is parent ion 100 Priority A Rule if annotation ion type is a 101 Priority C Rule if annotation ion type is c 102 Priority IM Rule if annotation is immonium ion 103 Priority Internal with P Rule if annotation is internal fragment and fragment sequence starts with P 104 Priority Internal without P Rule if annotation is internal fragment and fragment sequence starts without P 105 Priority Neutral loss Rule if annotation has neutral losses 106 Priority Enumeration end if there are more elements in the candidate list
then add annotation with neutral losses of H2O and C2H4O from T to the queue then add annotation with neutral loss of C3H6 from V to the queue then add annotation with neutral losses of C8H7N and C9H9N from W to the queue then add annotation with neutral loss of NH3 from immonium ion K then add annotation with neutral loss of H2O from C-Term then add annotation with neutral loss of NH3 from N-Term then add annotation with neutral loss of CO from parent ion then add annotation with neutral loss of CO from internal fragment then go to rule39 get next elements in the candidate list and initialize score of the annotation to 0 discard annotation and remove from candidate list and go to rule95 then increase score by 100 then increase score by 100 then increase score by 99 then increase score by 98 then increase score by 98 then increase score by 97 then increase score by 80 then increase score by 70 then decrease score by 5 times number of H2O, NH3 and CO and 30 times number of all other losses then add scored annotation to result list and go to rule39
else annotation cannot have neutral losses else annotation cannot have internal fragments
