Matrix-assisted laser desorption/ionization terminator

Two relatively new techniques, matrix assisted laser desorption ionization-lime of flight mass spectrometry (MALDI-TOF) and electrospray ionization (FS1), offer new possibilities for analysis of polymers with molecular weights in the tens of thousands. PS molecular weights as high as 1.5 million have been determined by MALDI-TOF. Recent reviews on the application of these techniques to synthetic polymers include those by Ilantoif54 and Nielen.555 The methods have been much used to provide evidence for initiation and termination mechanisms in various forms of living and controlled radical polymerization.550 Some examples of the application of MALDI-TOF and ESI in end group determination are provided in Table 3.12. The table is not intended to be a comprehensive survey. 

Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry has contributed remarkably to unravelling the termination and initiation steps of the styrene/CO copolymerisation catalysed by the highly active bis-chelated complex [Pd(bipy)2](Pp5)2 in TFE [40]. Chain-end group analysis of the material produced in the absence of BQ showed that the termination by P-H elimination is accompanied by three different initiators two palladium alkyls from Pd-H formed by reaction of the precursor with CO and water (a and b) and a palladium carboalkoxy species formed by reaction of the precursor with the fluorinated alcohol and CO (c) (Chart 7.4). The suppression of the chain-transfer by alcoholysis was proposed to be responsible for the enhanced stability of the palladium acyl intermediates and hence for the high molecular weight of the copolymers produced. 

Stevenson, T. I. Loo, J. A. Greis, K. D. 1998. Coupling capillary high-performance liquid chromatography to matrix-assisted laser desorption/ionization mass spectrometry and N-terminal sequencing of peptides via automated microblotting onto membrane substrates. Anal. Biochem., 262,99-109. 

Lazar AC, Kloczewiak MA, Mazsaroff I (2004), Matrix-assisted laser desorption/ ionization mass spectrometry for the evaluation of the C-terminal lysine distribution of a recombinant monoclonal antibody, Rapid Commun. Mass Spectrom. 18 239-244. 

Correct mass of purified proteins is determined using matrix-assisted laser desorption ionization-time of flight (MALDI-TOF). N-terminal sequencing and/ or LC-ES-MS can verify correct amino acid sequence. Folding of the recombinant protein should be compared to the natural counterpart by measuring CD-spectra. Furthermore, nuclear magnetic resonance (NMR) analysis can be performed to ensure the presence of tertiary structure, important for IgE binding activity (Neudecker et al. 2003). 

Thiede, B., Salnikow, J. and WittmannLiebold, B. (1997) C-terminal ladder sequencing by an approach combining chemical degradation with analysis by matrix-assisted-laser-desorption ionization mass spectrometry. Eur. J. Biochem., 244 (3), 750 4. 

There has been considerable debate over the reason why TEMPO does not work well for acrylates. CNRS researchers utilized NMR and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) techniques to look at the terminal end-groups on poly(n-butyl acrylate) produced using TEMPO mediation [19]. They found that most of the chains were terminated by co-unsaturation resulting from either elimination or O-H TEMP from the chain-end or disproportionation between TEMPO and the polyradical (Scheme 8.7). 

For peptide libraries, one solution to this problem uses matrix-assisted laser desorption ionization (MALDI) mass spectrometry to directly desorb and identify peptides from beads that were screened and found to be hits (33). This technique is called the termination synthesis approach. Because the peptide library compounds are analyzed directly, products with amino acid deletions or substitutions, side-reaction products, or incomplete deprotection are readily observed. Also, because there are no extra molecules used for chemical... 

Recombinant wild type hIL-ip, the K138C mutant and the K138C, R4A, L6A triple mutant (mutant 1) were isolated from the soluble fraction of E. coli lysates by ammonium sulfate fractionation and hydrophobic interaction chromatography. The purified proteins were characterized by SDS-PAGE, western blots, N-terminal sequence, size exclusion chromatography (SEC), isoelectric focusing (lEF), matrix assisted laser desorption ionization mass spectrometry (MALDI-MS), and electrospray mass spectrometry (ESMS). 

In a second step, the transesterification of the terminal ester group is possible by using 3-methylpentanediol. Owing to a steric effect, the reaction becomes selective, and it is possible to obtain a functionality very close to 2. The telechelic structure was evidenced by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) analysis. The telechelic oligomers were then reacted with 1,6-hexanediisocyanate to increase the molecular weight of the oligomers. 

DASH, dynamic allele-specific hybridization AS-PCR, allele-specific polymerase chain reaction AS-PE, allele-specific primer extension APEX, arrayed primer extension FP-TDI, fluorescence polarization template directed dye terminator incorporation MALDI-TOF-MS, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry OLA, oligonucleotide ligation assay RCA, ... 

Zammit, M. D., Davis, T. R, and Haddleton, D. M., Determination of the Propagation Rate Coefficient (kp) and Termination Mode in the Free-Radical Polymerization of Methyl Methacrylate, Employing Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry for Molecular Weight Distribution Anal5 is, Macromolecules, 29, 492, 1996. 

B. Kilster, T. J. P. Naven, and D. J. Harvey, Effect of the reducing-terminal substituents on the high energy collision-induced dissociation matrix-assisted laser desorption/ionization mass spectra of oligosaccharides. Rapid Commun. Mass Spectrom. 10, 1645-1651 (1996). 

Figure 7.3 K Ras4B C-terminal peptide was synthesized employing the trityl linker and after purification was reacted with a K-Ras thioester (K-Ras 1-174 MesNa) via EPL, yielding the semisynthetic K-Ras4B. Purity and identity of the purified protein were characterized by SDS-PAGE and MALDi-TOF. SDS-PAGE sodium dodecyl sulfate/polyacrylamide gel electrophoresis, MALDi-TOF matrix-assisted laser desorption ionization/time of flight.

Wang, D., Kalb, S.R., and Cotter, R.J. (2004) Improved procedures for N-terminal sulfonation of peptides for matrix-assisted laser desorption/ ionization post-source decay peptide sequendng. Rapid Commun. Mass Spectrom., 18, 95-102. 

Thiede, B. and Wittmann-Liebold, B. (2000) Analysis of RNA-protein cross-link sites by matrix-assisted laser desorption/ionization mass spectrometry and N-terminal microsequencing. Methods Enzymol., 318, 438-446. 

Derivatization procedures to facilitate de novo sequencing of lysine-terminated tryptic peptides using postsource decay matrix-assisted laser desorption/ionization... 

Fuchigami, T., Misumi, S., Takamnne, N., Takahashi, L, Takama, M. and Shoji, S., Acid-labile formylation of amino terminal prohne of human inrmnnodeficiency virus type 1 p24(gag) was found by proteomics using two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization-time-of-flightmass spectrometry. Biochem. Biophys. Res. Commun., 293, 1107-1113 (2002). 

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