Elucidation of Functionally Significant Structural Modifications
123
Table 8.1. Fragment Ions of Derivatives of the Peptide IMIKFNRL
Ion Type
m/z
Derivative
Byproduct
Asn6 Sequence
N~-Butyl-Asn6
Asp6 Sequence
Theory' Obs
Diff
Obs
Diff
Obs
Diff
Obs
Diff
C-terminal
z2
271.4
271.4
271.2
0.2
271.1
0.3
271.4
y2
288.4
288.5
0.1
288.3
0.1
288.3
0.1
288.5
0.1
z3
385.5
441.6
56.1
385.4
0.1
441.2
55.7
385.9
0.4
y3
402.5
458.6
56.1
402.3
0.2
458.3
55.8
403.5
1.0
z4
532.6
532.1
0.5
588.1
55.5
533.3
0.7
y4
549.7
605.9
56.2
549.1
0.6
605.0
55.3
550.2
0.5
z5
660.8
717.3
56.5
660.3
0.5
716.3
55.5
661.8
1.0
y5
677.8
734.3
56.5
677.3
0.5
733.2
55.4
678.8
1.0
z6
774
830.8
56.8
773.6
0.4
829.9
55.9
775.1
1.1
y6
791
848.1
57.1
791.1
0.1
846.9
55.9
792.1
1.1
z7
905.2
905.4
0.2
y7
922.2
978.6
56.4
921.4
0.8
977.6
55.4
922.9
0.7
z8
1018.3
1074.5
56.2
1017.6
0.7
1018.5
0.2
N-Terminal
al
86.2
86.2
86.2
86.1
0.1
86.2
cl
129.2
129.3
0.1
129.2
129.1
0.1
129.2
a2
217.4
217.5
0.1
217.3
0.1
217.3
0.1
217.5
0.1
b2
245.4
245.4
245.1
0.3
245.2
0.2
245.2
0.2
b3
358.5
358.7
0.2
358.3
0.2
358.3
0.2
357.5
0.8
c3
373.5
373.7
0.2
373.4
0.1
373.2
0.3
373.6
0.1
a4
458.7
458.6
0.1
458.3
0.4
b4
486.7
486.5
0.2
486.6
0.1
as
605.9
605.9
605.6
0.3
605.0
0.9
605.9
b5
633.9
634.4
0.5
633.3
0.6
633.7
0.2
633.8
0.1
b6
748
748.1
0.1
747.6
0.4
747.1
0.9
748.5
0.5
b7
904.2
960.6
56.4
903.8
0.4
959.8
55.6
905.4
1.2
b8
1017.3
1017.3
1073.4
56.1
1018.5
1.2
'Theoretical fragment ion masses are those calculated for the unmodified Asn6 version of the
sequence.
A further check on the validity of the assignments made to ions in the fragment ion spectrum of the immunologically active byproduct was performed by
examining the version of the sequence deliberately synthesized with N~-butylAsn at position 6. The ion series observed for this peptide were essentially the
same as those seen for the isolated byproduct except for variable observation of
z4, zS and bS fragments shifted by 56 Da relative to the unmodified sequence
(Fig. S.2C and S.3; Table 8.1). The potential N~-butyl-Asn immonium ion was
observed at m/z = 143.0. An ion corresponding to a b6 fragment was observed at
m/z = 747.1, which was a further indication that the corresponding ion seen for
the byproduct was produced by a combination of cleavage of the peptide chain
and cleavage of the butyl group attached to Asn6.
Analysis of fragments produced by the a-Asp6 form of the peptide (Fig. 8.3D;
Table 8.1) provided further support for the assignment of identities to ions in the
C-terminal series of the previous spectra. The aspartyl peptide produced a domi-