need help - (Nov/05/2008 )
would you please help me answering this question
In proteomics the most common approach is to digest the protein of interest using trypsin and analyse the resulting peptides by mass spectrometry. The masses are then searched against a protein database and the peptide masses are matched to the protein sequence. In most cases the sequence coverage (the number of matched amino-acid residues compared with the total number of amino-acids) can be quite low, say 20-50%, and is never 100%. A typical protein coverage is shown below:
Match to: HSP7C_HUMAN Score: 92 Expect: 1.2e-005
Heat shock cognate 71 kDa protein OS=Homo sapiens GN=HSPA8 PE=1 SV=1
Number of mass values matched: 12
Sequence Coverage: 26%
1 MSKGPAVGID LGTTYSCVGV FQHGKVEIIA NDQGNRTTPS YVAFTDTERL
51 IGDAAKNQVA MNPTNTVFDA KRLIGRRFDD AVVQSDMKHW PFMVVNDAGR
101 PKVQVEYKGE TKSFYPEEVS SMVLTKMKEI AEAYLGKTVT NAVVTVPAYF
151 NDSQRQATKD AGTIAGLNVL RIINEPTAAA IAYGLDKKVG AERNVLIFDL
201 GGGTFDVSIL TIEDGIFEVK STAGDTHLGG EDFDNRMVNH FIAEFKRKHK
251 KDISENKRAV RRLRTACERA KRTLSSSTQA SIEIDSLYEG IDFYTSITRA
301 RFEELNADLF RGTLDPVEKA LRDAKLDKSQ IHDIVLVGGS TRIPKIQKLL
351 QDFFNGKELN KSINPDEAVA YGAAVQAAIL SGDKSENVQD LLLLDVTPLS
401 LGIETAGGVM TVLIKRNTTI PTKQTQTFTT YSDNQPGVLI QVYEGERAMT
451 KDNNLLGKFE LTGIPPAPRG VPQIEVTFDI DANGILNVSA VDKSTGKENK
501 ITITNDKGRL SKEDIERMVQ EAEKYKAEDE KQRDKVSSKN SLESYAFNMK
551 ATVEDEKLQG KINDEDKQKI LDKCNEIINW LDKNQTAEKE EFEHQQKELE
601 KVCNPIITKL YQSAGGMPGG MPGGFPGGGA PPSGGASSGP TIEEVD
Peptides 1 and 5 are also cleaved by trypsin at an internal arginine (underlined).
Explain why it is not possible to obtain the full protein sequence (100% sequence coverage)?
In proteomics the most common approach is to digest the protein of interest using trypsin and analyse the resulting peptides by mass spectrometry. The masses are then searched against a protein database and the peptide masses are matched to the protein sequence. In most cases the sequence coverage (the number of matched amino-acid residues compared with the total number of amino-acids) can be quite low, say 20-50%, and is never 100%. A typical protein coverage is shown below:
Match to: HSP7C_HUMAN Score: 92 Expect: 1.2e-005
Heat shock cognate 71 kDa protein OS=Homo sapiens GN=HSPA8 PE=1 SV=1
Number of mass values matched: 12
Sequence Coverage: 26%
1 MSKGPAVGID LGTTYSCVGV FQHGKVEIIA NDQGNRTTPS YVAFTDTERL
51 IGDAAKNQVA MNPTNTVFDA KRLIGRRFDD AVVQSDMKHW PFMVVNDAGR
101 PKVQVEYKGE TKSFYPEEVS SMVLTKMKEI AEAYLGKTVT NAVVTVPAYF
151 NDSQRQATKD AGTIAGLNVL RIINEPTAAA IAYGLDKKVG AERNVLIFDL
201 GGGTFDVSIL TIEDGIFEVK STAGDTHLGG EDFDNRMVNH FIAEFKRKHK
251 KDISENKRAV RRLRTACERA KRTLSSSTQA SIEIDSLYEG IDFYTSITRA
301 RFEELNADLF RGTLDPVEKA LRDAKLDKSQ IHDIVLVGGS TRIPKIQKLL
351 QDFFNGKELN KSINPDEAVA YGAAVQAAIL SGDKSENVQD LLLLDVTPLS
401 LGIETAGGVM TVLIKRNTTI PTKQTQTFTT YSDNQPGVLI QVYEGERAMT
451 KDNNLLGKFE LTGIPPAPRG VPQIEVTFDI DANGILNVSA VDKSTGKENK
501 ITITNDKGRL SKEDIERMVQ EAEKYKAEDE KQRDKVSSKN SLESYAFNMK
551 ATVEDEKLQG KINDEDKQKI LDKCNEIINW LDKNQTAEKE EFEHQQKELE
601 KVCNPIITKL YQSAGGMPGG MPGGFPGGGA PPSGGASSGP TIEEVD
Peptides 1 and 5 are also cleaved by trypsin at an internal arginine (underlined).
Explain why it is not possible to obtain the full protein sequence (100% sequence coverage)?
What do you think so far? We can't/won't just answer it for you, it won't help you at all.