De novo Sequencing of Proteins With Mass Spectrometry
75
menting the lBO-isotope is not related to the peptide but is part of the chemical
background transmitted together with the peptide into the collision cell. The
subtracted spectrum emphasizes differences between the two tandem MS spectra
and makes the 16 0 isotope of the y3 ion apparent. This information could not
A
956.5
Ul 30
.... c
~
0
<.)
d
3 20
.c
' U;
940.5
s::
*
(1)
.... 10
*
. S
935
940
945
950
955
960
965
m/z, amu
B
25
956.5
y9
11
935
940
945
950
955
960
965
m/z, amu
Fig. 5.5. Details of the fragment spectrum of the peptide VAPAPAPAPEVQTK labelled to 50 % with
ISO. Panel A shows part of the native spectrum, panel B the corresponding part after subtraction of
the spectrum generated from the ISO labelled precursor. The y9 ion is clearly visible in the subtracted
spectrum but the 956.5 Da ion was retained as well even though it is obviously not a C-terminal fragment ion (panel A). This effect is caused by the limited perfection in aligning the two original data
sets. Therefore, it is important to consider the complete fragments spectrum as well, when reading
out the y ion series from the subtracted data set