Diagnostic NH and OH Vibrations for Oxazolone and Diketopiperazine Structures: b₂ from Protonated Triglycine

Abstract

We present infrared multiple photon dissociation (IRMPD) spectra in the hydrogen stretching region of the simplest b fragment, b 2 from protonated triglycine, contrasted to that of protonated cyclo(Gly-Gly). Both spectra confirm the presence of intense, diagnostic vibrations linked to the site of proton attachment. Protonated cyclo(Gly-Gly) serves as a reference spectrum for the diketopiperazine structure, showing a diagnostic O-H + stretch of the protonated carbonyl group at 3585 cm -1 . Conversely, b 2 from protonated triglycine exhibits a strong band at 3345 cm -1 , associated with the N-H stretching mode of the protonated oxazolone ring structure. Other weaker N-H stretches can also be discerned, such as the amino NH 2 and amide NH bands. These results demonstrate the usefulness of the hydrogen stretching region, and hence benchtop optical parametric oscillator/amplifier (OPO/A) set-ups, in making structural assignments of product ions in collision-induced dissociation (CID) of peptides.

Conformation-Specific Spectroscopy of Peptide Fragment Ions in a Low-Temperature Ion Trap

“…Although diketopiperazines are assumed to be the thermodynamically more stable isomers, oxazolones were found to prevail in many investigations [11,15,31,32,36,38,41,42,67]. The presence of histidine can apparently lower the high barrier to formation [66] of the two cis-amide bonds in diketopiperazines [39].…”

“…Room temperature IRMPD spectra in the XH stretching region were recently used to diagnose the protonation site in b 2 oxazolones [41,42], and structures with protonation at the oxazolone ring nitrogen have been reported as the major species observed in these investigations for the YI and GG b 2 ions, respectively. In contrast, under our conditions, we can largely exclude these in favor of species protonated on the N-terminal nitrogen atom for the YG b 2 ion.…”

“…Different tools, sometimes in combination, have been applied for obtaining information on the structures of peptide fragment ions, including theoretical calculations [9,10], multi-stage mass spectrometry (MS/MS, or MS n ) [3,4,[11][12][13][14][15][16][17][18][19][20][21][22][23][24][25][26][27][28], ion mobility spectrometry (IMS) [29,30], hydrogen-deuterium exchange (HDX) [22,31,32], and infrared multiple photon dissociation (IRMPD) spectroscopy [5,22,[31][32][33][34][35][36][37][38][39][40][41][42]. While spectroscopy is a particularly powerful way to obtain detailed information about the structures of gas phase molecules, approaches based on IRMPD feature certain drawbacks.…”

“…With increasing complexity of the fragment ion structures, support from other sources of information becomes necessary for unraveling complicated IRMPD spectra. This can be achieved by combining spectroscopy with other techniques [22,31,32] or by comparison of the spectra with chemically modified species in which cyclization is either induced or inhibited [36,38,39,42,44].…”

Disfavoring Macrocycle b Fragments by Constraining Torsional Freedom: The “Twisted” Case of QWFGLM b6

“…Thus, the oxazolone structure can be confirmed via the presence of a lactone-type C=O stretch band, which appears in the high-frequency region (1770-1950 cm -1 ) of the midinfrared spectrum [24,25]. Studies by several groups have shown that smaller b fragments, such as b 2 , typically form oxazolone structures [18,21,[26][27][28]. An exception to this rule is the b 2 sequence motif His-Ala, which mainly adopts a diketopiperazine structure [19].…”