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The peptide bond is the weakest point, so the protein will break into two pieces....

After collision with helium the peptide will break into characteristic fragments. The peptide bond is the weakest point, so the protein will break into two pieces. Expected fragments:. b-ions y-ions SLY K SL YK S LYK. Total 7 peaks. I. SLYK. LYK. K. YK. S. L. Y. m/z.

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The peptide bond is the weakest point, so the protein will break into two pieces....

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  1. After collision with helium the peptide will break into characteristic fragments The peptide bond is the weakest point, so the protein will break into two pieces....

  2. Expected fragments: b-ions y-ions SLY K SL YK S LYK Total 7 peaks I SLYK LYK K YK S L Y m/z From the fragment spectra the peptide sequence can be generated Example_1: Fragmentation of SLYK

  3. I SLYK SLY SL Y K m/z B-ions can help to identify the N-terminus of a peptide Expected fragments: b-ions y-ions SLY K SL YK S LYK Total 7 peaks

  4. I I SLY SLYK SLYK SL YK LYK K m/z m/z A completet spectra can be complex Expected fragments: b-ions y-ions SLY K SL YK S LYK Total 7 peaks

  5. How to interprete MS-spectra? Strategy #1 1. Try to find the C-terminal amino acid. 2. Try to find the next peak of the y-ion series in the spectra 3. Read the spectra from the tail to the head Strategy #2 • Find the peptide peak or the highest peak MW of one series. • Find the next peak in the series. • When the sequence is done, try to figure out in which way to read it. Combined Strategy • Find the b2/a2 pair • Read the b-ion series as far as you can • Read the y-ion series. • Try to match them together

  6. Example:

  7. Peptide C-terminal amino acid Δ163 Δ233 Δ128 Δ113 Δ113 Δ113 Δ101 Δ129 Δ87

  8. List of masses of amino acid and modifications m/z A=71, P=97, V=99, I/L=113, N=114, D=115, M=131, H=137, F=147, K=129/147, Q=128, S=87 Y=163, E= 129 T=101 R=175 phosphorylation = +80 ICPL= +86/92

  9. Sequence: YQKSTE(I/L)(I/L)(I/L)R 215 Y R 173 ICPL- K (13C) Q K(12C) I/L I/L T E S I/L

  10. A, Overexpression and enhanced activity of the epidermal growth factor receptor (EGFR) is often observed in human cancers. EGFR is a tyrosine phosphotransferase (kinase) and abnormal EGFR-kinase activity is related to poor prognosis. Upon binding of the ligand EGFR undergoes autophosphorylation (self phosphorylation), an important step in EGF signal transduction. Figure 1 shows the fragment spectra of the peptide (PDYQQDFFPK) generated from isolated EGFR. Please confirmthe predicted sequence and mark the phosphorylation site. Figure 1: Fragment spectrum. (m/z A=71, P=97, V=99, I,=113, N=114, D=115, M=131, H=137, F=147, K=129/147, Q=128, Y=163, phosphorylation = +80) B, A pharmaceutical company wants to produce inhibitors against EGFR-kinase for cancer therapy. Please explain briefly how mass spectrometry can help to identify an inhibitor compound.

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