Activation of large lons in FT‐ICR mass spectrometry

Ultraviolet photodissociation of peptides followed by mass analysis has several desirable advantages relative to other methods, yet it has not found widespread use due to several limitations. One shortcoming is the inefficiency with which peptides absorb in the ultraviolet. This issue has a simple solution and can be circumvented by the attachment of noncovalent adducts that contain appropriate chromophores. Subsequent photoactivation of the chromophore leads to vibrational excitation of the complex and eventually to fragmentation of the peptide. Herein, the energetics that control the efficiency of this process are examined as a function of the characteristics of both the peptide and the noncovalently attached chromophore. Fragmentation efficiency decreases with increasing peptide size and is also constrained by the binding energy of the noncovalent adduct. The optimum chromophore should have excellent absorption at the excitation wavelength and a low luminescence quantum yield. It is demonstrated that a naphthyl based 18-crown-6 adduct is ideally suited for attaching to a variety peptides and fragmenting them following absorption of 266 nm light. [7][8][9][10][11]. Though none of these techniques is perfect, each has characteristic strengths and weaknesses. For example CID is easily employed in the source region of any instrument with atmospheric pressure ionization. CID is also easily accommodated elsewhere in an instrument as long as high vacuum is not required. In contrast, IRMPD is well suited for collisionless environments. In this regard, CID and IRMPD provide complementary capabilities. Furthermore, since both methods fragment ions by the stepwise addition of small amounts of energy, the results obtained are frequently quite similar.However, CID and IRMPD are both limited by the fact that energy deposition occurs over a relatively long time scale, typically milliseconds to seconds. In contrast, ultraviolet photodissociation (UVPD) occurs on a much shorter time scale and can yield fragment ions characteristic of CID or IRMPD. However, the major limitation of UVPD is that the precursor ion must contain a suitable chromophore. For peptides, UVPD requires either short wavelength lasers (less than 200 nm) or peptides rich in tyrosine, tryptophan, and phenylalanine that can absorb at longer wavelengths. Therefore, UVPD is either limited to a small subset of all peptides or constrained by undesirable technical challenges which are encountered when working in the vacuum ultraviolet. Thus, widespread implementation of UVPD for peptide fragmentation has been to date somewhat limited.However, Brodbelt and coworkers have recently described a method to circumvent the requirement for aromatic residues or short wavelength lasers [12]. In this work, a noncovalently attached chromophore was used to absorb energy from a photon and transfer it to a peptide, causing the peptide to fragment. 18-crown-6 ether (18C6), which preferentially associates with lysine residues via the formation of three hydrogen bonds, was used for...

Section: Resultsmentioning

confidence: 99%

Rapid peptide fragmentation without electrons, collisions, infrared radiation, or native chromophores

Yeh

Sun

Meneses

et al. 2009

“…In low-energy collisionactivated dissociation (CAD) and infrared multiphoton dissociation (IRMPD), the peptide ion is heated in a multi-step process [7,8]. The model of the mobile proton can rationalize the observed fragments [9]; the transfer of a proton weakens locally the peptide bond, facilitating its fragmentation.…”

mentioning

confidence: 99%

Comparative dissociation of peptide polyanions by electron impact and photo-induced electron detachment

Larraillet

Vorobyev

Brunet

et al. 2010

We compare product-ion mass spectra produced by electron detachment dissociation (EDD) and electron photodetachment dissociation (EPD) of multi-deprotonated peptides on a Fourier transform and a linear ion trap mass spectrometer, respectively. Both methods, EDD and EPD, involve the electron emission-induced formation of a radical oxidized species from a multi-deprotonated precursor peptide. Product-ion mass spectra display mainly fragment ions resulting from backbone cleavages of C ␣ -C bond ruptures yielding a and x ions. Fragment ions originating from N-C ␣ backbone bond cleavages are also observed, in particular by EPD. Although EDD and EPD methods involve the generation of a charge-reduced radical anion intermediate by electron emission, the product ion abundance distributions are drastically different. Both processes seem to be triggered by the location and the recombination of radicals (both neutral and cation radicals). Therefore, EPD product ions are predominantly formed near tryptophan and histidine residues, whereas in EDD the negative charge solvation sites on the backbone seem to be the most favorable for the nearby bond dissociation. (J Am Soc Mass Spectrom 2010, 21, 670 -680) © 2010 Published by Elsevier Inc. on behalf of American Society for Mass Spectrometry O ne of the main advantages of mass spectrometry for peptide and protein structure analysis is its ability to give valuable information from very low amounts of samples, e.g., primary structure (amino acids sequence), number and location of disulfide bridges, identification, and sometimes characterization of post-translational modifications. To provide this information, various activation techniques are used to fragment peptide or protein ions to yield structurespecific product ions complementary to accurate molecular mass measurements [1][2][3][4][5][6].Different methods are available nowadays to excite and fragment biomolecular ions. In low-energy collisionactivated dissociation (CAD) and infrared multiphoton dissociation (IRMPD), the peptide ion is heated in a multi-step process [7,8]. The model of the mobile proton can rationalize the observed fragments [9]; the transfer of a proton weakens locally the peptide bond, facilitating its fragmentation. Since the fragmentation occurs after heating and vibrational energy redistribution, structural rearrangements are in competition with fragmentation pathways. In addition and in complement to slow heating methods, reactions of polypeptide ions with electrons and small radical ions have become a very useful tool for peptide structural analysis. For instance, electron capture dissociation (ECD) [6,10] and electron-transfer dissociation (ETD) [11][12][13] may initiate bond breaking in peptide and protein polycations presumably faster than energy redistribution over all degrees of freedom. Less than a decade ago, an ionelectron interaction-based fragmentation technique for peptide and protein polyanions termed electron detachment dissociation (EDD) was proposed [14 -16]. More recently, an ion-ion int...

mentioning

confidence: 99%

“…The first example of IRMPD in a quadrupole ion trap mass spectrometer used a multipass optical system to increase the path length of the laser and overlap with the ion cloud [1]. In Fourier transform ion cyclotron resonance mass spectrometers collisional cooling is not an issue, but because of the large magnetron radius of the ions the laser is commonly left unfocused to maximize laser overlap [18]; irradiation times are commonly up to one order of magnitude longer than for IRMPD in a quadrupole ion trap mass spectrometer. The work reported here shows that focusing the laser for IRMPD in a quadrupole ion trap mass spectrometer is the most simple and effective means of increasing fragmentation efficiency.…”

mentioning

confidence: 99%

Improving IRMPD in a quadrupole ion trap

Newsome

Glish

2009

A focused laser is used to make infrared multiphoton photodissociation (IRMPD) more efficient in a quadrupole ion trap mass spectrometer. Efficient (up to 100%) dissociation at the standard operating pressure of 1 ϫ 10 Ϫ3 Torr can be achieved without any supplemental ion activation and with shorter irradiation times. The axial amplitudes of trapped ion clouds are measured using laser tomography. Laser flux on the ion cloud is increased six times by focusing the laser so that the beam waist approximates the ion cloud size. Unmodified peptide ions from 200 Da to 3 kDa are completely dissociated in 2.5-10 ms at a bath gas pressure of 3.3 ϫ 10 Ϫ4 Torr and in 3-25 ms at 1.0 ϫ 10 Ϫ3 Torr. Sequential dissociation of product ions is increased by focusing the laser and by operating at an increased bath gas pressure to minimize the size of the ion cloud. nfrared multiphoton photodissociation (IRMPD) has been used for tandem mass spectrometry in a quadrupole ion trap mass spectrometer [1][2][3][4][5][6][7] and a Fourier transform ion cyclotron resonance mass spectrometer [8 -11] as an alternative to collision-induced dissociation (CID). Only one m/z ion is selected for excitation in conventional CID, limiting sequential dissociation of product ions into smaller mass product ions. Conversely, in IRMPD all parent and product ions are irradiated simultaneously with a collimated, IR laser providing the capability to generate abundant small mass product ions. IRMPD is typically performed in a quadrupole ion trap mass spectrometer at a smaller low mass cut-off (LMCO) than CID, allowing the observation of a broader m/z range of product ions [2,4], and a higher MS/MS efficiency,where E MS/MS is MS/MS efficiency, F i is the abundance of each product ion, and P 0 is the initial abundance of the parent ion before irradiation. IRMPD is performed in a Fourier transform ion cyclotron resonance mass spectrometer to eliminate the need for a CID target gas to be leaked into and pumped out of the ICR cell [10,11]. The sensitivity of a quadrupole ion trap mass spectrometer is optimized with a bath gas pressure around 1.0 ϫ 10 Ϫ3 Torr. Bath gas collisions reduce the kinetic energy of trapped ions and minimize the size of the ion cloud [12]. However, collisions also remove internal energy gained from IR absorption, necessitating longer periods of irradiation for dissociation and decreasing fragmentation efficiency [13],where E F is fragmentation efficiency, F i is the abundance of each product ion, and P is the abundance of the parent ion remaining after irradiation. IRMPD in a quadrupole ion trap mass spectrometer often is performed at 1 ϫ 10 Ϫ5 to 3 ϫ 10 Ϫ4 Torr to decrease collisional cooling of parent ion internal energy [2-7], [13] and achieve dissociation with shorter irradiation times. However, this reduced bath gas pressure is less effective for trapping ions during injection and reduces sensitivity.Several methods have been developed to increase IRMPD fragmentation efficiency by combining IRMPD with another form of ion activation an...