252CF plasma desorption mass spectrometry of a polycyclic peptide antibiotic, Nisin (original) (raw)
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Journal of Mass Spectrometry, 2006
reflectron instrument fitted with a gas collision cell were used. For optimum comparability of data from different IT instruments, the CID conditions were standardized and only singly charged precursor ions were considered. Additionally, HE-CID data obtained from the TOF-based instrument were acquired and compared with LE-CID data from ITs. Major differences between trap-based and TOF-based CID data are that the latter data set lacks abundant additional loss of small neutrals (e.g. ammonia, water) but contains product ions down to the immonium-ion-type region, thereby allowing the detection of even single amino-acid (even unusual amino acids) substitutions. For several polypeptide antibiotics, mass spectrometric as well as tandem mass spectrometric data are shown and discussed for the first time, and some yet undescribed minor components are also reported. De novo sequencing of unusually linked minor components of (e.g. cyclic) polypeptides is practically impossible without knowledge of the exact structure and fragmentation behavior of the major components. Finally, the described standardized CID condition constitutes a basic prerequisite for creating a searchable, annotated MS n -database of bioactive compounds. The applied desorption/ionization techniques showed no significant influence on the type of product ions (neglecting relative abundances of product ions formed) observed, and therefore the type of analyzer connected with the CID process mainly determines the type of fragment ions.
Biological Mass Spectrometry, 1988
Molecular ion yields and spectrum quality in plasma desorption mass spectrometry of peptides and proteins have been studied as a function of the method of sample preparation. Parameters like matrix preparation, pH of the sample solution, method of applying the sample on the matrix and sample quantity have been studied. The best matrix was found to be nitrocellulose prepared by electrospraying. Various techniques for applying the sample result in different sensitivity. The highest sensitivity is obtained using the spin-drying technique, which results in highquality spectra with 1-2 pmol of trypsin (mol. wt 23,463). The results obtained using different sample prep aration techniques are discussed and directions for an optimal procedure are given.
Journal of the American Society for Mass Spectrometry, 2003
Desfuroylceftiofur (DFC) is a bioactive -lactam antibiotic metabolite that has a free thiol group. Previous experiments have shown release of DFC from plasma extracts after addition of a disulfide reducing agent, suggesting that DFC may be bound to plasma and tissue proteins through disulfide bonds. We have reacted DFC with [Arg 8 ]-vasopressin (which has one disulfide bond) and bovine insulin (which has three disulfide bonds) and analyzed the reaction products by use of electron capture dissociation Fourier transform ion cyclotron resonance mass spectrometry (ECD FT-ICR MS), which has previously shown preferential cleavage of disulfide bonds. We observe cleavage of DFC from vasopressin and insulin during ECD, suggesting that DFC is indeed bound to peptides and proteins through disulfide bonds. Specifically, we observed dissociative loss of one, as well as two, DFC species during ECD of [vasopressin ϩ 2(DFC-H) ϩ 2H] 2ϩ from a single electron capture event. Loss of two DFCs could arise from either consecutive or simultaneous loss, but in any case implies a gas phase disulfide exchange step. ECD of [insulin ϩ DFC ϩ 4H] 4ϩ shows preferential dissociative loss of DFC. Combined with HPLC, ECD FT-ICR-MS may be an efficient screening method for detection of drug-biomolecule binding.
Applications of plasma desorption mass spectrometry in peptide and protein chemistry
Biomedical & environmental mass spectrometry
Molecular weight determination by plasma desorption (PD) mass spectrometry has been used in the different stages of protein sequence determination. It has been found to be a very valuable addition to the traditional techniques. For analysis in biotechnological development and production PD mass spectrometry meets the require ments for speed, precision and mass range. The amount of information can be increased by combining 'wet' biochemical procedures with analysis by PD mass spectrometry.
Analytical Biochemistry, 1998
The lantibiotic nisin and some of its variants and degbeen proposed more recently (2), ranging from the proradation products have been characterized, using a 9.4duction of natural cheese to uses as a therapeutic agent T Fourier transform ion cyclotron resonance mass specin combination with chelators. Thus, there is a considtrometer and electrospray ionization. The abundances erable interest in understanding the structure-funcof all products in the sample (i.e., major component, varition relationships in this molecule and in modifying its ants, degradation products, and adducts) have been structure to enhance its performance as an antibiotic. measured quantitatively. The mass resolution obtained Protein engineering strategies have been developed to in the electrospray ionisation mass spectra was approxiachieve these ends (3, 4), but development in parallel mately 100,000 over the measured range. The resulting of sophisticated methods for confirming structures of mass accuracy, better than 0.7 ppm (or within 0.001 Da) nisin, its degradation products, its variants, precurallowed the molecular masses and in many cases chemisors, and new mutants produced is highly desirable.
International Journal of Mass Spectrometry, 2004
Although electron capture dissociation (ECD) offers many advantages for structural elucidation, a fundamental understanding of all possible processes following electron capture is necessary if ECD is to succeed in the characterization of unknowns. Many biologically active compounds have non-standard structures, e.g, N-alkylation, branching, cyclization, and ester linkages. Here we report ECD of cyclodepsipeptides (valinomycin and beauvericin), including N-methylated structures (beauvericin), branched peptides (AcA 3 K(G 3 )A 3 -NH 2 and A 3 K(G 3 )A 3 -NH 2 ), and oligomers of ε-amino acids (ε-peptides) (Ac(Ahx) 6 K and (Ahx) 6 K) to establish the behavior of such non-standard structures. ECD of cyclodepsipeptides yielded numerous backbone fragments but no charge-reduced species, consistent with a radical cascade mechanism. ECD of ε-peptides resulted in a • and y fragments only, suggesting that the N-C␣ c/z • fragmentation channel is impeded in those structures. ECD of branched peptides resulted in complex fragmentation patterns, characterized by the presence of the immonium related m ion from the modified residue.
Chemical-ionization mass spectrometry of .BETA.-lactam antibiotics
The Journal of Antibiotics, 1975
Chemical-ionization (CI) mass spectra are described for methyl esters of eight clinically significant penicillins and their breakdown products. These substances give spectra with very few fragment ions and contain easily discernible protonated molecule ions. The main cleavage reaction is postulated to involve a retro 2 + 2 DIELS-ALDER-type fragmentation of the 13-lactam ring liberating one fragment (m/e=174) that is characteristic of the penicillin nucleus and a second fragment that is molecule specific, as it contains the elements of the side chain. The other fragment ions, though interesting, are of minor intensity. The free acids, on the other hand, fragment more extensively because of their relative instability and lack of volatility. These spectra resemble electron impact spectra more closely and, though they encode more structural information, are less reproducible from run to run. The ease with which the esters can be made and the relative simplicity of their CI mass spectra make this method significant for the identification and characterization of 3-lactam antibiotics.
Journal of Chromatography A, 2005
A rapid method has been developed for the quantification of the prototypic cyclotide kalata B1 in water and plasma utilizing matrix-assisted laser desorption ionisation time-of-flight (MALDI-TOF) mass spectrometry. The unusual structure of the cyclotides means that they do not ionise as readily as linear peptides and as a result of their low ionisation efficiency, traditional LC/MS analyses were not able to reach the levels of detection required for the quantification of cyclotides in plasma for pharmacokinetic studies. MALDI-TOF-MS analysis showed linearity (R 2 > 0.99) in the concentration range 0.05-10 g/mL with a limit of detection of 0.05 g/mL (9 fmol) in plasma. This paper highlights the applicability of MALDI-TOF mass spectrometry for the rapid and sensitive quantification of peptides in biological samples without the need for extensive extraction procedures.