9780521516358book.pdf

residue and post-translational modification to N-terminal trimethylalanine and

dimethylproline also occur. In the case of recombinant proteins over-expressed in

E. coli, the initiator residueN-formyl methionine is often incompletely removed.

All these modifications leave the N-terminal residue without a free proton on the

alpha nitrogen and Edman chemistry cannot proceed. Mass spectrometry has there-

fore been essential for their correct structural identification. The protein sequencing

x 3 y 3 z 3 x 2 y 2 z 2 x 1 y 1 z 1

H 2 N C

R 1

H

O

N

R 2

H

C

O

N

H

R 3 R 4

H

C

O

N

H

C

H

CO 2 H

H

a 1 b 1 c 1 a 2 b 2 c 2 a 3 b 3 c 3

CCC

(a)

+

b 1 1207.61

b 2 1060.54

b 3 959.50

b 4 862.44

b 5 765.39

b 6 708.37

b 7 580.31

b 8 509.27

b 9 438.24

b 10 275.17

b 11

130.05 E FTPPGQAAYQK y 11

277.12 EF TPPGQAAYQK y 10

378.17 EFT PPGQAAYQK y 9

475.22 EFTP PGQAAYQK y 8

572.27 EFTPP GQAAYQK y 7

629.29 EFTPPG QAAYQK y 6

757.35 EFTPPGQ AAYQK y 5

828.39 EFTPPGQA AYQK y 4

899.43 EFTPPGQAA YQK y 3

1062.49 EFTPPGQAAY QK y 2

1190.55 EFTPPGQAAYQ K y 1 147.11

+

(b)

b 4

475.22

b 7

757.35

b 8

b 828.39

2

277.12

b 3

378.17

b 6

629.29 b^9

899.43

b 11

1190.55

b 5

572.27

b 10

1062.49

y 9

959.50

y 8

862.44

y 7

765.39

y 6

708.37

y 5 y 10 1060.54

580.31

y 4

y438.24 (^3) 509.27
y 11 1207.61
y 2
275.17
y 1 147.11
b 1
130.1
(M+H)–1336.65
0 100 1000
Fig. 9.22Peptide fragment ion nomenclature and tandem MS spectrum of a peptide. (a) Charge may be
retained by either the N- or C-terminal fragment, resulting in the a, b and c series of ions or x, y and z series
respectively. Ions in the b and y series frequently predominate. Corresponding neutral fragments are of course
not detected. (b) The sequence of the peptide from a mutant haemoglobin is: EFTPPGQAAYQK. The figure
shows the tandem mass spectrum from collision-induced dissociation of the doubly charged (Mþ2H)^2 þ
precursor,m/z¼668.3. Cleavage at each peptide bond results in the b or y ions when the positive charge is
retained by the fragment containing the N- or C-terminus of the peptide respectively (see inset).
383 9.5 Structural information by tandem mass spectrometry