Binding of Divalent Metal Ions with Deprotonated Peptides: Do Gas-Phase Anions Parallel the Condensed Phase?

Chelation complexes of the histidine-containing tripeptides HisAlaAla, AlaHisAla, and AlaAlaHis with Ni(II) and Cu(II) having a -1 net charge are characterized in the gas phase by infrared multiple-photon dissociation (IRMPD) spectroscopy and density functional theory calculations. We address the question of whether the gas-phase complexes carry over characteristics from the corresponding condensed-phase species. We focus particularly on three aspects of their structure: (i) square-planar chelation by the deprotonated amide nitrogens around the metal ion (low-spin for the Ni case), (ii) metal-ion coordination of the imidazole side chain nitrogen, and (iii) the exceptional preference for metal-ion chelation by peptides with His in the third position from the N-terminus, as in the amino terminal Cu and Ni (ATCUN) motif. We find that square-planar binding around the metal ion, involving bonds to both deprotonated backbone nitrogens, one of the carboxylate oxygens and the N-terminal nitrogen, is the dominant binding motif for all three isomers. In contrast to the condensed-phase behavior, the dominant mode of binding for all three isomers does not involve the imidazole side chain, which is instead placed outside the coordination zone. Only for the AlaAlaHis isomer, the imidazole-bound structure is also detected as a minority population, as identified from a distinctive short-wavelength IR absorption. The observation that this conformation exists only for AlaAlaHis correlates with condensed-phase behavior at neutral-to-basic pH, in the sense that the isomer with His in the third position is exceptionally disposed to metal ion chelation by four nitrogen atoms (4N) when compared with the other isomers. These results also emphasize the divergence between the conformational stabilities in the gas phase and in solution or crystalline environments: in the gas phase, direct metal binding of the imidazole is overall less favorable than the alternative of a remote imidazole that can act as an intramolecular H-bond donor enhancing the gas-phase stability.

Infrared laser dissociation of single megadalton polymer ions in a gated electrostatic ion trap: the added value of statistical analysis of individual events.

In this study, we report the unimolecular dissociation mechanism of megadalton SO3-containing poly(2-acrylamido-2-methyl-1-propanesulfonic acid) (PAMPS) polymer cations and anions with the aid of infrared multiphoton dissociation coupled to charge detection ion trap mass spectrometry. A gated electrostatic ion trap ("Benner trap") is used to store and detect single gaseous polymer ions generated by positive and negative polarity in an electrospray ionization source. The trapped ions are then fragmented due to the sequential absorption of multiple infrared photons produced from a continuous-wave CO2 laser. Several fragmentation pathways having distinct signatures are observed. Highly charged parent ions characteristically adopt a distinctive "stair-case" pattern (assigned to the "fission" process) whereas low charge species take on a "funnel like" shape (assigned to the "evaporation" process). Also, the log-log plot of the dissociation rate constants as a function of laser intensity between PAMPS positive and negative ions is significantly different.

Water Microsolvation Can Switch the Binding Mode of Ni(II) with Small Peptides.

Ni(II) ions can be caged by surrounding peptide ligands in two basic binding patterns: the "iminol" (IM) binding pattern, where chelation occurs by deprotonated amide nitrogens, or the charge-solvated (CS) binding pattern, where chelation occurs by amide carbonyl oxygens. Gas-phase observation may clarify the factors affecting this choice in solution and in peptide and protein matrices. Infrared spectroscopic determination of gas-phase structures shows here how microsolvation by just one water molecule switches the balance of this choice from IM to CS for the Ni2+Gly3 complex, in contrast with the always-CS structure of the Ni2+Gly4 complex. Quantum-chemical calculations indicate that CS complexation is even more favored in the aqueous limit. Considering gas-phase conditions as comparable to low-pH solutions can reconcile this prediction with the common observation of IM-type binding in solutions at higher pH. This is likely the first gas-phase observation of solvation-induced IM-to-CS transition in oligopeptide complexes with doubly charged transition-metal ions.

Complexes of Ni(ii) and Cu(ii) with small peptides: deciding whether to deprotonate.

The observed variety of metal-ion complexation sites offered by peptides reflects a basic tension between charge solvation of the ion by Lewis-basic chelating groups versus amide nitrogen deprotonation and formation of metal-nitrogen bonds. Gas-phase models of metal-ion coordination can illuminate the factors governing this choice in condensed-phase proteins and enzymes. Here, structures of gas-phase complexes of Ni(ii) and Cu(ii) with tri- and tetra-peptide ligands are mapped out using a combination of Infrared Multiple Photon Dissociation (IRMPD) spectroscopy and density functional theory (DFT) computations. The two binding modes give distinctive IRMPD signatures, particularly in the diagnostic region 1500-1550 cm-1. Previous observations have suggested that Ni(ii) complexes preferentially show the iminol rearrangement pattern (Im) giving low-spin square-planar geometries with metal-ion bonds to deprotonated amide nitrogens. In contrast, alkaline earth metal ion complexes prefer amide carbonyl oxygens chelating the metal ion with pyramidal geometry (charge-solvation, CS). Surprisingly, it is shown here that the Gly4 complexes are CS bound, in contrast with the expectation of Im binding. It is suggested that CS binding is actually a normal Ni(ii) and Cu(ii) binding mode to simple peptides lacking participating side chains. Three factors are suggested to influence the choice between CS and Im binding patterns: (1) presence of an accessible side-chain Lewis-basic proton interaction site (FGGF, FGG and HAA complexes); (2) short chain length of the peptide leading to a shortage of accessible carbonyl oxygen sites for CS binding, (AAA, FGG and HAA complexes); (3) outright deprotonation of the ligand giving net negatively charged Im[Ni2+(Gly4-3H+)]- and Im[Ni2+(Ala3-3H+)]- complexes, which have a triply-deprotonated ligand. IRMPD spectra of [Cu2+Gly4]2+ and [Cu2+(Gly4-3H+)]- complexes suggest that their structures are similar to their Ni2+ analogs.

Divalent Metal-Ion Complexes with Dipeptide Ligands Having Phe and His Side-Chain Anchors: Effects of Sequence, Metal Ion, and Anchor.

Conformational preferences have been surveyed for divalent metal cation complexes with the dipeptide ligands AlaPhe, PheAla, GlyHis, and HisGly. Density functional theory results for a full set of complexes are presented, and previous experimental infrared spectra, supplemented by a number of newly recorded spectra obtained with infrared multiple photon dissociation spectroscopy, provide experimental verification of the preferred conformations in most cases. The overall structural features of these complexes are shown, and attention is given to comparisons involving peptide sequence, nature of the metal ion, and nature of the side-chain anchor. A regular progression is observed as a function of binding strength, whereby the weakly binding metal ions (Ba(2+) to Ca(2+)) transition from carboxylate zwitterion (ZW) binding to charge-solvated (CS) binding, while the stronger binding metal ions (Ca(2+) to Mg(2+) to Ni(2+)) transition from CS binding to metal-ion-backbone binding (Iminol) by direct metal-nitrogen bonds to the deprotonated amide nitrogens. Two new sequence-dependent reversals are found between ZW and CS binding modes, such that Ba(2+) and Ca(2+) prefer ZW binding in the GlyHis case but prefer CS binding in the HisGly case. The overall binding strength for a given metal ion is not strongly dependent on the sequence, but the histidine peptides are significantly more strongly bound (by 50-100 kJ mol(-1)) than the phenylalanine peptides.

Spectroscopy of metal-ion complexes with peptide-related ligands.

With new experimental tools and techniques developing rapidly, spectroscopic approaches to characterizing gas-phase metal ion complexes have emerged as a lively area of current research, with particular emphasis on structural and conformational information. The present review gives detailed attention to the metal-ion complexes of amino acids (and simple derivatives), much of whose study has focused on the question of charge-solvation vs salt-bridge modes of complexation. Alkali metal ions have been most frequently examined, but work with other metal ions is discussed to the extent to which they have been studied. The majority of work has been with simple cationic metal ion complexes, while recent excursions into deprotonated complexes, anionic complexes, and dimer complexes are also of interest. Interest is growing in complexes of small peptides, which are discussed both in the context of possible zwitterion formation as a charge-solvation alternative, and of the alternative metal-ion bond formation to amide nitrogens in structures involving iminol tautomerization. The small amount of work on complexes of large peptides and proteins is considered, as are the structural consequences of solvation of the gas-phase complexes. Spectroscopy in the visible/UV wavelength region has seen less attention than the IR region for structure determination of gas-phase metal-ion complexes; the state of this field is briefly reviewed.

Metal ion complexes with HisGly: comparison with PhePhe and PheGly.

Gas-phase complexes of five metal ions with the dipeptide HisGly have been characterized by DFT computations and by infrared multiple photon dissociation spectroscopy (IRMPD) using the free electron laser FELIX. Fine agreement is found in all five cases between the predicted IR spectral features of the lowest energy structures and the observed IRMPD spectra in the diagnostic region 1500-1800 cm(-1), and the agreement is largely satisfactory at longer wavelengths from 1000 to 1500 cm(-1). Weak-binding metal ions (K(+), Ba(2+), and Ca(2+)) predominantly adopt the charge-solvated (CS) mode of chelation involving both carbonyl oxygens, an imidazole nitrogen of the histidine side chain, and possibly the amino nitrogen. Complexes with Mg(2+) and Ni(2+) are found to adopt iminol (Im) binding, involving the deprotonated amide nitrogen, with tetradentate chelation. This tetradentate coordination of Ni(II) is the preferred binding mode in the gas phase, against the expectation under condensed-phase conditions that such binding would be sterically unfavorable and overshadowed by other outcomes such as metal ion hydration and formation of dimeric complexes. The HisGly results are compared with corresponding results for the PheAla, PheGly, and PhePhe ligands, and parallel behavior is seen for the dipeptides with N-terminal Phe versus His residues. An exception is the different chelation pattern determined for PhePhe versus HisGly, reflecting the intercalation-type cation binding pocket of the PhePhe ligand. The complexes group into three well-defined spectroscopic patterns: nickel and magnesium, calcium and barium, and potassium. Factors leading to differentiation of these distinct spectroscopic categories are (1) differing propensities for choosing the iminol binding pattern, and (2) single versus double charge on the metal center. Nickel and magnesium ions show similar gas-phase binding behavior, contrasting with their quite different patterns of peptide interaction in condensed phases.

Metal cation binding to gas-phase pentaalanine: divalent ions restructure the complex.

Ion-neutral complexes of pentaalalanine with several singly- and doubly charged metal ions are examined using conformation analysis by infrared multiple photon dissociation (IRMPD) spectroscopy and density functional theory (DFT) computations. The infrared spectroscopy in the 1500-1800 cm(-1) region is found to be conformationally informative; in particular, the frequency of the C═O stretching mode of the terminal carboxyl group is diagnostic for hydrogen bonding of the terminal hydroxyl. The doubly charged alkaline earth metal ions (Ca(2+) and Ba(2+)) enforce a highly structured chelation shell around the metal ion, with six strongly bound Lewis-basic chelation sites, and no hydroxyl hydrogen bonding. With the more weakly binding alkali metal ions (Na(+), K(+), and Cs(+)), structures with intramolecular hydrogen bonds are more favorable, leading to dominance of conformations with lower degrees of metal ion chelation. The favored coordination mode correlates with ionic charge and binding strength but is not related to the ionic radius of the metal ion.

Encapsulation of metal cations by the PhePhe ligand: a cation-π ion cage.

Structures and binding thermochemistry are investigated for protonated PhePhe and for complexes of PhePhe with the alkaline-earth ions Ba(2+) and Ca(2+), the alkali-metal ions Li(+), Na(+), K(+), and Cs(+), and the transition-metal ion Ag(+). The two neighboring aromatic side chains open the possibility of a novel encapsulation motif of the metal ion in a double cation-π configuration, which is found to be realized for the alkaline-earth complexes and, in a variant form, for the Ag(+) complex. Experimentally, complexes are formed by electrospray ionization, trapped in an FT-ICR mass spectrometer, and characterized by infrared multiple photon dissociation (IRMPD) spectroscopy using the free electron laser FELIX. Interpretation is assisted by thermochemical and IR spectral calculations using density functional theory (DFT). The IRMPD spectrum of protonated PhePhe is reproduced with good fidelity by the calculated spectrum of the most stable conformation, although the additional presence of the secondmost stable conformation is not excluded. All metal-ion complexes have charge-solvated binding modes, with zwitterion (salt bridge) forms being much less stable. The amide oxygen always coordinates to the metal ion, as well as at least one phenyl ring (cation-π interaction). At least one additional chelation site is always occupied, which may be either the amino nitrogen or the carboxy carbonyl oxygen. The alkaline-earth complexes prefer a highly compact caged structure with both phenyl rings providing cation-π stabilization in a "sandwich" configuration (OORR chelation). The alkali-metal complexes prefer open-cage structures with only one cation-π interaction, except perhaps Cs(+). The Ag(+) complex shows a unique preference for the closed-cage amino-bound NORR structure. Ligand-driven perturbations of normal-mode frequencies are generally found to correlate linearly with metal-ion binding energy.

When do molecular bowls encapsulate metal cations?

Curved carbon π surfaces have chemical and physical properties suitable for exploitation for chemical microencapsulation and the self-assembly of nanoscale materials. Advances will greatly benefit from more understanding of their host-guest interactions with guests such as metal cations. Here, quantitative predictions are made for the binding of metal cations to three prototypical surfaces using density functional theory calculations: the buckybowls C(20)H(10), C(30)H(10), and C(40)H(10). The focus was on finding the most favorable binding sites, assessing whether binding is more favorable inside or outside the bowl, and exploring factors influencing the binding site preference. Classes of cations studied included small and large monocations and cations with multiple charges: Na(+), Cs(+), NH(4)(+), Ba(+), Ba(2+), and La(3+). Factors found to favor inside binding were large ion size and high ion charge, suggesting that polarization interactions as well as short-range interactions are important in determining the preferred binding sites inside and outside these buckybowls. Unlike monocations, which at best have only a weak tendency toward encapsulation, the multiply charged cations Ba(2+) and La(3+) were found to have a strong driving force toward containment inside the bowls. Coulomb potentials were found to favor cation binding on the outside surface of the bowls, but cation microsolvation through polarization interactions presents a compensating factor that can tip the balance in favor of encapsulation. Knowledge of these factors will be a valuable tool in the design of nanocontainers and the diverse architecture possible with these structural elements.

Chirality-induced conformational preferences in peptide-metal ion binding revealed by IR spectroscopy.

Chirality reversal of a residue in a peptide can change its mode of binding to a metal ion, as shown here experimentally by gas-phase IR spectroscopy of peptide-metal ion complexes. The binding conformations of Li(+), Na(+), and H(+) with the LL and DL stereoisomers of PhePhe were compared through IR ion spectroscopy using the FELIX free-electron laser. For the DL isomer, both Li(+) and Na(+) exclusively coordinate to the amide O atom, the carboxyl O atom, and one of the aromatic rings (the OOR conformation), while for the LL isomer, a mixture of the OOR and NOR conformations was found. The stereochemically induced change in conformation is shown to reflect the strength of an NH···π interaction remote from the metal ion site. Protonated PhePhe shows no stereochemically induced variation in binding geometry.

Cationized phenylalanine conformations characterized by IRMPD and computation for singly and doubly charged ions.

Electrospray ionization produces phenylalanine (Phe) complexes of the alkali metal ion series, plus Ag(+) and Ba(2+). Infrared multiple photon dissociation (IRMPD) spectroscopy using the FELIX free electron laser light source is used to characterize the conformations of the ions, in conjunction with density functional theory (DFT) calculations giving thermochemical information and computed infrared spectra for likely candidate conformations. For complexes of small, singly charged ions (Li(+), Na(+), K(+) and Ag(+)) a single tridentate, charge-solvated conformational theme (N/O/Ring) binding amino nitrogen, carbonyl oxygen and the aromatic ring to the metal ion accounts for all the observations. The larger alkalis Rb(+) and Cs(+) show clear spectroscopic evidence of mixed populations, containing substantial fractions of both tridentate and also bidentate chelation. For Rb(+) the bidentate fraction is assigned as the (O/Ring) chelation pattern, while for Cs(+) a mixture of (O/Ring) and (O/O) chelation patterns seems likely. All of the smaller ions with high positive charge density have a clear preference for cation-π interaction with the side-chain aromatic ring, but for the larger ions Rb(+) and particularly Cs(+) this interaction becomes sufficiently weak to allow conformations having the metal ion remote from the π system. The Ba(2+) complex is unique in showing clear evidence of a major fraction of salt-bridge (zwitterionic) ions along with charge-solvated conformations. Plots of the frequency shifts of the two highly perturbed ligand vibrational modes (C[double bond, length as m-dash]O stretch and NH(2) frustrated inversion) give good linear correlations with the binding energy of the metal to the ligand.

Ion spectroscopy: where did it come from; where is it now; and where is it going?

The ASMS conference on ion spectroscopy brought together at Asilomar on October 16-20, 2009 a large group of mass spectrometrists working in the area of ion spectroscopy. In this introduction to the field, we provide a brief history, its current state, and where it is going. Ion spectroscopy of intermediate size molecules began with photoelectron spectroscopy in the 1960s, while electronic spectroscopy of ions using the photodissociation "action spectroscopic" mode became active in the next decade. These approaches remained for many years the main source of information about ionization energies, electronic states, and electronic transitions of ions. In recent years, high-resolution laser techniques coupled with pulsed field ionization and sample cooling in molecular beams have provided high precision ionization energies and vibrational frequencies of small to intermediate sized molecules, including a number of radicals. More recently, optical parametric oscillator (OPO) IR lasers and free electron lasers have been developed and employed to record the IR spectra of molecular ions in either molecular beams or ion traps. These results, in combination with theoretical ab initio molecular orbital (MO) methods, are providing unprecedented structural and energetic information about gas-phase ions.

Peptide length, steric effects, and ion solvation govern zwitterion stabilization in barium-chelated di- and tripeptides.

Infrared multiple-photon dissociation (IRMPD) spectroscopy has given infrared spectra of complexes of di- and tripeptides (AlaAla, AlaAlaAla, AlaPhe, PheAla) with singly and doubly charged metal ions (K(+), Ca(2+), Sr(2+), and Ba(2+)). The switch between charge-solvated (CS) and salt-bridged zwitterion (SB) conformations is displayed through highly diagnostic features in the mid-infrared. Systematic trends are found correlating with the length of the peptide chain (tripeptides favoring CS conformations), metal ion size (larger metals favoring SB conformations), metal ion charge (doubly charged ions favoring SB conformations), and sterically available Lewis-basic side-chain interactions with the metal ion (for example a cation-pi interaction with Ba(2+) stabilizes CS for PheAla but not for AlaPhe). The principle is that CS conformations are favored for small metal ions with high charge density and extensive microsolvation of the charge by Lewis-basic groups, especially amide carbonyls; SB conformations are favored by metal ions of high charge but low charge density, which are better stabilized by salt-bridge Coulomb interactions.

Conformation switching in gas-phase complexes of histidine with alkaline earth ions.

Infrared multiple photon dissociation spectroscopy of gas-phase doubly charged alkaline earth complexes of histidine reveals a transition from dominance of the zwitterion (salt bridge, SB) conformation with Ba2+ to substantial presence of the canonical (charge-solvated, CS) conformation with Ca2+. This result is a clear illustration of the importance of metal-ion size in governing the delicate balance between these two modes of complexation of gas-phase amino acids. The two conformational motifs are clearly distinguished by characteristic spectral features, confirmed by density functional theory simulated IR spectra of the low-energy conformers. As a further illustration of histidine complexation possibilities, the spectrum of the Na+His complex shows purely CS character and emphasizes the greater tendency toward SB character induced by the higher charge in the alkaline earth complexes. Calculation of the complete series of alkaline earth/histidine complexes confirms the increasing stability of the SB conformations relative to CS with increasing metal ion size, as well as showing that among SB conformations the most highly chelated conformation (SB3) is favored for small metals, whereas the most extended conformation (SB1) is favored for large metals. A decomposition of the binding thermochemistry shows that these thermochemical trends versus metal-ion size are due to differences in electrostatic binding energies, with relatively little contribution from the deformation and rearrangement energy costs of distorting the ligand framework.

Dimeric complexes of tryptophan with M2+ metal ions.

IRMPD spectroscopy using the FELIX free electron laser and a Fourier transform ICR mass spectrometer was used to characterize the structures of electrosprayed dimer complexes M(2+)Trp(2) of tryptophan with a series of eight doubly charged metal ions, including alkaline earths Ca, Sr, and Ba, and transition metals Zn, Cd, Mn, Co, and Ni. With the support of DFT thermochemical calculations, at least three different structural motifs were distinguished spectroscopically, depending critically on the nature of the metal ion. The spectral signatures of a ligand in the charge-solvated (CS) configuration, namely peaks near 1730 and 1150 cm(-1), were prominent in all the spectra, and it was clear that all the dimer complexes contain at least one CS ligand. The spectra indicated that the second ligand is zwitterionic (ZW) for all complexes except the Ni case, with the second ligand having an extended binding geometry with smaller metals but showing some admixture of a compact chelated geometry with larger alkaline earths. It was concluded that these dimer complexes have a mixed configuration of ligands, denoted CS/ZW. The Ni(2+)Trp(2) complex is exceptional, with the spectroscopy and the thermochemical calculation both indicating a CS/CS configuration of ligands. This geometry appears to correlate with the exceptionally small size and high binding strength of the Ni(2+) cation. The complex CdClTrp(1+) was also obtained and gave a clear spectrum showing a CS ligand configuration. The presence of a CS ligand in all the dimeric complexes of the 2+ metals is an interesting contrast with the monomer complex Ba(2+)Trp, in which the ligand is ZW.

Alkali metal complexes of the dipeptides PheAla and AlaPhe: IRMPD spectroscopy.

Complexes of PheAla and AlaPhe with alkali metal ions Na(+) and K(+) are generated by electrospray ionization, isolated in the Fourier-transform ion cyclotron resonance (FT-ICR) ion trapping mass spectrometer, and investigated by infrared multiple-photon dissociation (IRMPD) using light from the FELIX free electron laser over the mid-infrared range from 500 to 1900 cm(-1). Insight into structural features of the complexes is gained by comparing the obtained spectra with predicted spectra and relative free energies obtained from DFT calculations for candidate conformers. Combining spectroscopic and energetic results establishes that the metal ion is always chelated by the amide carbonyl oxygen, whilst the C-terminal hydroxyl does not complex the metal ion and is in the endo conformation. It is also likely that the aromatic ring of Phe always chelates the metal ion in a cation-pi binding configuration. Along with the amide CO and ring chelation sites, a third Lewis-basic group almost certainly chelates the metal ion, giving a threefold chelation geometry. This third site may be either the C-terminal carbonyl oxygen, or the N-terminal amino nitrogen. From the spectroscopic and computational evidence, a slight preference is given to the carbonyl group, in an RO(a)O(t) chelation pattern, but coordination by the amino group is almost equally likely (particularly for K(+)PheAla) in an RO(a)N(t) chelation pattern, and either of these conformations, or a mixture of them, would be consistent with the present evidence. (R represents the pi ring site, O(a) the amide oxygen, O(t) the terminal carbonyl oxygen, and N(t) the terminal nitrogen.) The spectroscopic findings are in better agreement with the MPW1PW91 DFT functional calculations of the thermochemistry compared with the B3LYP functional, which seems to underestimate the importance of the cation-pi interaction.