Anal. Chem. 2007, 79, 2901-2910
Electron Capture Dissociation of Oligosaccharides Ionized with Alkali, Alkaline Earth, and Transition Metals Julie T. Adamson and Kristina Håkansson*
Department of Chemistry, University of Michigan, 930 North University Avenue, Ann Arbor, Michigan 48109-1055
We extend the application of electron capture dissociation (ECD) (which requires at least two charges) to oligosaccharides without basic functionalities by utilizing alkali, alkaline earth, and transition metals (Na+, K+, Ca2+, Ba2+, Mg2+, Mn2+, Co2+, and Zn2+) as charge carriers in electrospray ionization. Both linear and branched oligosaccharides were examined, including maltoheptoase, p-lactoN-hexaose, and an N-linked glycan from human r1-acid glycoprotein. For comparison, infrared multiphoton dissociation (IRMPD) was also applied to all oligosaccharide species. We show that, for certain metal-adducted oligosaccharides, particularly maltoheptaose, cross-ring cleavage, which can provide saccharide linkage information, is the dominant fragmentation pathway in ECD. By contrast, glycosidic cleavages dominate in IRMPD although cross-ring fragmentation was also observed to varying degrees depending on metal ion type. The branched N-linked glycan did not fragment as easily following ECD compared to the linear oligosaccharides, presumably due to intramolecular noncovalent interactions. However, this limitation was partially overcome with a combined ECD/ IRMPD approach (activated ion ECD). For all metaladducted oligosaccharides, complementary structural information was obtained with ECD as compared to IRMPD. Our results demonstrate that ECD of metaladducted oligosaccharides is a valuable tool for structural characterization of oligosaccharides. Oligosaccharide-containing biomolecules, known as glycoconjugates, are highly diverse and prevalent in biological systems. The role of glycoconjugates in nature is extensive,1 ranging from protein folding2 to immune system response.3 The multiple functions of glycoconjugates are largely due to the increased degree of complexity oligosaccharides impart to these biomolecules. Oligosaccharides may exist as several isomeric forms with diverse linkages and, unlike other biomolecules, can form highly branched structures. Complete structural characterization of oligosaccharides requires information regarding linkage, sequence, branching, and anomeric configuration (a term that refers * To whom correspondence should be addressed. E-mail:
[email protected]. Tel: (734) 615-0570. Fax: (734) 647 4865. (1) Varki, A. Glycobiology 1993, 3, 97-130. (2) Parodi, A. J. Annu. Rev. Biochem. 2000, 69, 60-93. (3) Rudd, P. M.; Elliott, T.; Cresswell, P.; Wilson, I. A.; Dwek, R. A. Science 2001, 291, 2370-2376. 10.1021/ac0621423 CCC: $37.00 Published on Web 03/01/2007
© 2007 American Chemical Society
to the configuration, R or β, of the glycosidic bond of a sugar). Structural characterization of the oligosaccharide portion of a glycoconjugate is often accomplished by releasing the sugar from the biomolecule. It is frequently necessary to utilize a wide range of analytical methodologies in order to fully characterize these diverse and structurally complex molecules. Mass spectrometry is an important tool for oligosaccharide characterization and offers high sensitivity and minimum sample requirements. Tandem mass spectrometry (MSn) has been employed extensively for oligosaccharide structural analysis.4-13 In particular, Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) offers several advantages for oligosaccharide analysis,14 including high mass accuracy, ultrahigh resolution, and the availability of several tandem mass spectrometric techniques. Oligosaccharides undergo two main types of fragmentation. The predominant fragmentation pathway for protonated oligosaccharides is generally glycosidic cleavage, which occurs between monosaccharide units and provides information regarding saccharide sequence and branching. However, valuable information regarding sugar linkage can be gained from cross-ring cleavages, which are less prevalent than glycosidic cleavages. Several factors are known to affect the degree of oligosaccharide fragmentation and the extent of glycosidic versus cross-ring cleavage. These factors include variables such as the ionizing cation, the lifetime of the ion prior to detection, and the energy deposited into the ion. Oligosaccharides ionized with alkali, alkaline earth, and transition metals often fragment to yield more cross-ring cleavages (4) Carr, S. A.; Reinhold, V. N.; Green, B. N.; Haas, J. R. Biomed. Mass Spectrom. 1985, 12, 288-295. (5) Mu ¨ ller, D. R.; Domon, B.; Blum, W.; Raschdorf, F.; Richter, W. J. Biomed. Environ. Mass Spectrom. 1988, 15, 441-446. (6) Domon, B.; Mu ¨ ller, D. R.; Richter, W. J. Org. Mass Spectrom. 1989, 24, 357-359. (7) Domon, B.; Mu ¨ ller, D. R.; Richter, W. J. Biomed. Environ. Mass Spectrom. 1990, 19, 390-392. (8) Laine, R. A.; Pamidimukkala, K. M.; French, A. D.; Hall, R. W.; Abbas, S. A.; Jain, R. K.; Matta, K. L. J. Am. Chem. Soc. 1988, 110, 6931-6939. (9) Domon, B.; Mu ¨ ller, D. R.; Richter, W. J. Int. J. Mass Spectrom. Ion Processes 1990, 100, 301-311. (10) Gillece-Castro, B. L.; Burlingame, A. L. Methods Enzymol. 1990, 193, 689712. (11) Lemoine, J.; Strecker, G.; Leroy, Y.; Fournet, B.; Ricart, G. Carbohydr. Res. 1991, 221, 209-217. (12) Harvey, D. J. Mass Spectrom. Rev. 1999, 18, 349-451. (13) Zaia, J. Mass Spectrom. Rev. 2004, 23, 161-227. (14) Park, Y.; Lebrilla, C. B. Mass Spectrom. Rev. 2005, 24, 232-264.
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compared to their protonated counterparts.15-27 These fragmentation patterns are highly dependent on the specific metal adduct chosen and also the degree of oligosaccharide branching.22 In addition, fragmentation of deprotonated oligosaccharides may also result in additional cross-ring fragmentation.10,28-30 Conventionally, tandem mass spectrometry is accomplished with low-energy collision-activated dissociation (CAD) and results in mostly glycosidic cleavages. However, several alternative fragmentation techniques have been utilized for oligosaccharides including high-energy CAD,11,31-33 infrared multiphoton dissociation (IRMPD),34-36 electron capture dissociation (ECD),37 and 157nm photodissociation.38 High-energy CAD and 157-nm photodissociation of oligosaccharides results in more extensive cross-ring fragmentation. IRMPD and low-energy CAD are both low-energy vibrational excitation techniques. However, Lebrilla and coworkers have shown that the fragmentation efficiency in IRMPD is greater than that in CAD for large oligosaccharides.36 In addition, ECD has been applied to protonated chitooligosaccharides and yielded primarily glycosidic cleavages corresponding to B- and C-type ions. ECD is based on the dissociative recombination of polycationic molecules with low-energy electrons (
