Without more info, here's my educated guess at an explanation: that peptide is not a fully tryptic peptide in the proteins you list as it is preceded by a methionine residue. For isoform P06396-2, the preceding methionine is the first residue of the sequence and I'm assuming you did a Comet search with the "clip_nterm_methionine <http://comet-ms.sourceforge.net/parameters/parameters_201801/clip_nterm_methionine.php>" parameter set to "1". With that start methionine removed, this allowed that peptide to be fully tryptic in that particular isoform hence only that isoform is listed as a matched protein. The enzyme context does matter with RefreshParser; it's simply not a peptide string match.
On Wed, Jun 5, 2019 at 11:15 AM Vidya Venkatraman <vidya.venk...@gmail.com> wrote: > Hi Everyone, > > I am trying to run xinteract with the -D option so that RefreshParser can > use the fasta database to find all proteins corresponding to identified > peptides. > > This is the command I am using: > xinteract -OARPwd -dDECOY_ -PPM -nR > -D/scratch/venkatramanv/iphronesis/reference_files/Databases/UP_Human_Rev_Can+Iso_20190410_DECOY.fasta > -N/scratch/venkatramanv/iphronesis/platform_workspace/jobs/SJOB2122/peptideprophet/2122_PDAY_DDA_Isoforms.comet.interact.pep.xml > *.comet.pep.xml > > Despite using the -D database option, i noticed that some peptides in the > interact.pep.xml are matched uniquely to a specific isoform (even though > that peptide is not proteotypic or unique to any isoform). > > For example: > > <spectrum_query > spectrum="XL_150506_IDA_PDAY_F8_proteinpilot.08799.08799.2" > start_scan="8799" end_scan="8799" precursor_neutral_mass="1096.592165" > assumed_charge="2" index="20868" retention_time_sec="2254.3"> > <search_result> > <search_hit hit_rank="1" peptide="VVEHPEFLK" peptide_prev_aa="M" > peptide_next_aa="A" protein="sp|P06396-2|GELS_HUMAN" num_tot_proteins="1" > num_matched_ions="11" tot_num_ions="16" calc_neutral_pep_mass="1096.591696" > massdiff="0.000470" num_tol_term="1" num_missed_cleavages="0" > num_matched_peptides="6184"> > <search_score name="xcorr" value="2.205"/> > <search_score name="deltacn" value="0.575"/> > <search_score name="deltacnstar" value="0.000"/> > <search_score name="spscore" value="350.4"/> > <search_score name="sprank" value="1"/> > <search_score name="expect" value="4.94E-03"/> > <analysis_result analysis="peptideprophet"> > <peptideprophet_result probability="1.0000" > all_ntt_prob="(0.0000,1.0000,1.0000)"> > <search_score_summary> > <parameter name="fval" value="5.5754"/> > <parameter name="ntt" value="1"/> > <parameter name="nmc" value="0"/> > <parameter name="massd" value="0.429"/> > <parameter name="isomassd" value="0"/> > <parameter name="RT" value="1372.21"/> > <parameter name="RT_score" value="0.02"/> > </search_score_summary> > </peptideprophet_result> > </analysis_result> > </search_hit> > </search_result> > </spectrum_query> > > See attached the sequence alignment of all isoforms of GELS_HUMAN, you can > see that this peptide matches the canonical form as well as all isoforms > hence the num_tot_proteins should be = "4" and protein should be =" > sp|P06396|GELS_HUMAN" with <alternative_protein protein=" > sp|P06396-2|GELS_HUMAN"/>, > <alternative_protein protein="sp|P06396-3|GELS_HUMAN"/> , > <alternative_protein protein="sp|P06396-4|GELS_HUMAN"/> > > I am happy to share all the comet.pep.xml & interact.pep.xml files along > with the fasta database via Box folder. > > Can someone please help me understand if this is expected behavior? if so, > how i can fix this? > > Regards > Vidya > > -- > You received this message because you are subscribed to the Google Groups > "spctools-discuss" group. > To unsubscribe from this group and stop receiving emails from it, send an > email to spctools-discuss+unsubscr...@googlegroups.com. > To post to this group, send email to spctools-discuss@googlegroups.com. > Visit this group at https://groups.google.com/group/spctools-discuss. > To view this discussion on the web visit > https://groups.google.com/d/msgid/spctools-discuss/6b292366-55e1-4fe2-9994-ea9fff3d97a9%40googlegroups.com > <https://groups.google.com/d/msgid/spctools-discuss/6b292366-55e1-4fe2-9994-ea9fff3d97a9%40googlegroups.com?utm_medium=email&utm_source=footer> > . > For more options, visit https://groups.google.com/d/optout. > -- You received this message because you are subscribed to the Google Groups "spctools-discuss" group. To unsubscribe from this group and stop receiving emails from it, send an email to spctools-discuss+unsubscr...@googlegroups.com. To post to this group, send email to spctools-discuss@googlegroups.com. Visit this group at https://groups.google.com/group/spctools-discuss. To view this discussion on the web visit https://groups.google.com/d/msgid/spctools-discuss/CAJqD6ENwEimtBu3twOwHUPzSQ_m0Q2bzNpqiYcghbpaoRqPUkA%40mail.gmail.com. For more options, visit https://groups.google.com/d/optout.
