Characterization of noncovalent protein-ligand complexes and associated enzyme intermediates of GlcNAc-6-<i>O</i>-sulfotransferase by electrospray ionization FT-ICR mass spectrometry

Yu, Yonghao; Kirkup, Colleen E.; Pi, Na; Leary, Julie A.

doi:10.1016/j.jasms.2004.06.002

Cited by 38 publications

(38 citation statements)

References 31 publications

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“…Absolute K a values, in the 10 3 -10 7 M −1 range, have been determined for a variety of protein-ligand complexes, including antibody-antigen, lectin-carbohydrate, enzymeinhibitor complexes, and shown to be in reasonable agreement with values determined by other analytical methods [4][5][6][7][8][9][10]. To date, however, there have been few reported examples of the application of ESI-MS to characterize protein-hydrophobic ligand binding [11][12][13][14] and, to our knowledge, no reports of absolute affinity measurements using the direct ESI-MS assay.…”

Section: Introductionsupporting

confidence: 57%

Quantifying Protein-Fatty Acid Interactions Using Electrospray Ionization Mass Spectrometry

Liu

Kitova

Klassen

2011

J. Am. Soc. Mass Spectrom.

104

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The application of the direct electrospray ionization mass spectrometry (ESI-MS) assay to quantify interactions between bovine β-lactoglobulin (Lg) and a series of fatty acids (FA), CH 3 (CH 2 ) x COOH, where x=6 (caprylic acid, CpA), 8 (capric acid, CA), 10 (lauric acid, LA), 12 (myristic acid, MA), 14 (palmitic acid, PA) and 16 (stearic acid, SA), is described. Control ESI-MS binding measurements performed on the Lg-PA interaction revealed that both the protonated and deprotonated gas phase ions of the (Lg + PA) complex are prone to dissociate in the ion source, which leads to artificially small association constants (K a ). The addition of imidazole, a stabilizing solution additive, at high concentration (10 mM) increased the relative abundance of (Lg + PA) complex measured by ESI-MS in both positive and negative ion modes. The K a value measured in negative ion mode and using sampling conditions that minimize insource dissociation is in good agreement with a value determined using a competitive fluorescence assay. The K a values measured by ESI-MS for the Lg interactions with MA and SA are also consistent with values expected based on the fluorescence measurements. However, the K a values measured using optimal sampling conditions in positive ion mode are significantly lower than those measured in negative ion mode for all of the FAs investigated. It is concluded that the protonated gaseous ions of the (Lg + FA) complexes are kinetically less stable than the deprotonated ions. In-source dissociation was significant for the complexes of Lg with the shorter FAs (CpA, CA, and LA) in both modes and, in the case of CpA, no binding could be detected by ESI-MS. The affinities of Lg for CpA, CA, and LA determined using the reference ligand ESI-MS assay, a method for quantifying labile protein-ligand complexes that are prone to in-source dissociation, were found to be in good agreement with reported values.

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Section: Introductionsupporting

confidence: 57%

Quantifying Protein-Fatty Acid Interactions Using Electrospray Ionization Mass Spectrometry

Liu

Kitova

Klassen

2011

J. Am. Soc. Mass Spectrom.

104

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“…Generation of the noncovalent complex ions was performed as previously described (30). Briefly, samples were infused into the mass spectrometer at 1 l/min using a syringe pump (Harvard Apparatus, Holliston, MA).…”

Section: Methodsmentioning

confidence: 99%

“…Wild type MCP-1 (40 M) was mixed with 200 M of the octasaccharide library (30), and the solution was loaded onto an ultrafiltration unit. After washing the retentate with 200 mM NH 4 OAc, the resulting solution was analyzed by ESI-FTICR mass spectrometry.…”

Section: Esi-fticr Analysis Of Wt Mcp-1-mentioning

confidence: 99%

“…The binding stoichiometry as well as the molecular formula of the oligosaccharide binders can be calculated from the masses of the protein/oligosaccharide noncovalent complex using electrospray ionization-Fourier transform ion cyclotron resonance (ESI-FTICR) mass spectrometry. This is possible given the fact that many structural features, including stoichiometry, ligand binding domain, and even tertiary geometry of solution noncovalent complexes can be preserved during the transition from solution to gas phase (27)(28)(29)(30)(31). Confirmation of the complexed oligosaccharides detected in the noncovalent complex was verified using hydrophobic trapping and elution.…”

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confidence: 99%

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Chemokine-Glycosaminoglycan Binding

Sweeney

Saad

et al. 2005

Journal of Biological Chemistry

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Glycosaminoglycans (GAGs) have recently been demonstrated to be required for the in vivo activity of several chemokines. Minimally, the interaction is thought to provide a mechanism for retention at the site of secretion and the formation of chemokine gradients that provide directional cues for receptor bearing cells, particularly in the presence of shear forces. Thus, a key issue will be to determine the sequence and structure of the GAGs that bind to specific chemokines. Herein, we describe a mass spectrometry assay that was developed to detect protein-oligosaccharide noncovalent complexes, in this case chemokine-GAG interactions, and to select for high affinity GAGs. The process is facilitated by the ability of electrospray ionization to transfer the intact noncovalent complexes from solution into the gas phase. The elemental composition as well as the binding stoichiometry can be calculated from the mass of the complex. Ligands of the chemokine receptor, CCR2 (MCP-1/ CCL2, MCP-2/CCL8, MCP-3/CCL7, MCP-4/CCL13, and Eotaxin/ CCL11), and the CCR10 ligand CTACK/CCL27 were screened against a small, highly sulfated, heparin oligosaccharide library with limited structural variation. The results revealed heparin octasaccharides with 11 and 12 sulfates as binders. Oligomerization of some chemokines was observed upon GAG binding, whereas in other instances only the monomeric noncovalent complex was identified. The results indicate that, in contrast to the apparent redundancy in the chemokine system, where several chemokines bind and activate the same receptor, these chemokines could be differentiated into two groups based on the stoichiometry of their complexes with the heparin oligosaccharides.

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“…It is widely accepted that, under the appropriate conditions, the mass spectra of protein complexes can be used to determine the stoichiometry of interacting subunits [12] as well as the binding of substrates to active sites [13]. Electrospray mass spectrometry has gained increasing popularity in the characterization of self-assembled, highly charged supramolecular structures [3, 6 -10, 14 -16].…”

mentioning

confidence: 99%

Characterization of self-assembled supramolecular [Ga₄L₆] host-guest complexes by electrospray ionization mass spectrometry

Andersen

Seeber

Fiedler

et al. 2006

J. Am. Soc. Mass Spectrom.

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Self-assembled supramolecular host-guest complexes have been characterized by electrospray ionization mass spectrometry. The spectra obtained by use of a Q-TOF instrument equipped with a Z-spray ion source show primarily the 3-and 4-charge states of the assemblies. The assemblies have the general formula [guest ʚ Ga 4 L 6 ] 11Ϫ where L represents the chelating bidentate catechol ligand 1,5-bis(2=,3=-dihydroxy-benzamido)naphthalene and guests are tetramethyl ammonium (Me 4 N ϩ ), tetraethyl ammonium (Et 4 N ϩ ), tetra-n-propyl ammonium (Pr 4 N ϩ ) and decamethylcobaltocenium (Cp* 2 Co ϩ ) cations. For the first time, the mass spectrum of the empty assembly [Ga 4 L 6 ] 12Ϫ is reported. This article also reports that provided the electrospray ion source is capable of preserving noncovalent interactions, it is possible to observe host-guest complexes containing both weak binding guests as well as sterically demanding guests in the mass spectra. The present data suggest that electrospray mass spectrometry is a powerful tool for characterization of supramolecular host-guest complexes. ( I n the area of supramolecular chemistry, metal-ligand interactions have been exploited to design highly symmetric closed shell structures that selfassemble from simple metal and ligand components. The formation of discrete molecular architectures relies on labile coordinative bonds that allow for self-correction and exclusive formation of the thermodynamic product. A great variety of self-assembled structures of different molecular sizes and shapes is present in the literature [1][2][3][4][5]. Here, we focus on contributions to this field made by Raymond and coworkers [2, 6 -10]. The common theme in these metal-ligand assemblies is the presence of chelating bidentate catechol amide ligands, which bear certain geometric constraints upon coordination to octahedral metal centers resulting in M 4 L 6 coordination tetrahedra (Figure 1). The complexes are highly charged, rendering them soluble in polar solvents such as water and methanol, yet contain hydrophobic cavities of variable sizes. The presence of these hydrophobic cavities affords rich host-guest chemistry, enabling encapsulation of monocationic guest molecules [2, 6 -8]. Several analytical methods are necessary to efficiently characterize these host-guest complexes. 1 H-NMR spectroscopy shows remarkable upfield shifts in the proton signals of encapsulated guests within the cavities [2]. However, since the structures exhibit high point symmetry and the metal components are often paramagnetic, complete characterization by NMR is not always possible. Although solid-state structures may, in the best of cases, be determined by single crystal X-ray crystallography, intrinsically labile metal-ligand interactions do not always allow for unambiguous characterization of solution state structures. Additionally, mass spectrometry is more sensitive than NMR spectroscopy and allows analysis of lower concentrations, thus observing species with solubility properties insufficient for NMR measureme...

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Characterization of noncovalent protein-ligand complexes and associated enzyme intermediates of GlcNAc-6-O-sulfotransferase by electrospray ionization FT-ICR mass spectrometry

Cited by 38 publications

References 31 publications

Quantifying Protein-Fatty Acid Interactions Using Electrospray Ionization Mass Spectrometry

Quantifying Protein-Fatty Acid Interactions Using Electrospray Ionization Mass Spectrometry

Chemokine-Glycosaminoglycan Binding

Characterization of self-assembled supramolecular [Ga₄L₆] host-guest complexes by electrospray ionization mass spectrometry

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Characterization of noncovalent protein-ligand complexes and associated enzyme intermediates of GlcNAc-6-O-sulfotransferase by electrospray ionization FT-ICR mass spectrometry

Cited by 38 publications

References 31 publications

Quantifying Protein-Fatty Acid Interactions Using Electrospray Ionization Mass Spectrometry

Quantifying Protein-Fatty Acid Interactions Using Electrospray Ionization Mass Spectrometry

Chemokine-Glycosaminoglycan Binding

Characterization of self-assembled supramolecular [Ga4L6] host-guest complexes by electrospray ionization mass spectrometry

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Characterization of self-assembled supramolecular [Ga₄L₆] host-guest complexes by electrospray ionization mass spectrometry