Approaches to the characterization of membrane channel proteins (porins) by UV MALDI-MS

Schnaible, Volker; Michels, Jenny; Zeth, Kornelius; Freigang, Jörg; Welte, Wolfram; Bühler, Stefan; Glocker, Michael O.; Przybylski, Michael

doi:10.1016/s0168-1176(97)00226-7

Cited by 28 publications

(34 citation statements)

References 32 publications

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“…Therefore, the matrix and analyte molecules must be protected from separation in the crystallization step in the conventional solvent-based MALDI sample preparation. Certainly, minimization of separation or maximization of homogeneity can be introduced by optimization of the solvent-based sample preparation [7,[30][31][32][33][34][35][36][37]. Thus, similar polarities of the analyte and matrix molecules lead inevitably to reduction of separation during the crystallization based on our past experiences in solvent-based approaches.…”

Section: Direct Solvent-free Maldi-tof-ms Applying 25-dhbmentioning

confidence: 99%

“…In any solvent-based sample preparation, a separation of the analyte and matrix has to be prevented. Optimizing the conventional solvent-based MALDI sample preparation conditions can circumvent this intrinsic problem and make the MS analysis a great success; a few are cited [30][31][32][33][34][35][36][37]. In case of inhomogeneities, increasing the laser power during the measurement can enforce a sufficient desorption/ionization process, so that the analyte becomes accessible [14,25].…”

Section: Introductionmentioning

confidence: 99%

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Investigations of theoretical principles for MALDI-MS derived from solvent-free sample preparation

Trimpin

Räder

Müllen

2006

International Journal of Mass Spectrometry

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Section: Direct Solvent-free Maldi-tof-ms Applying 25-dhbmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Investigations of theoretical principles for MALDI-MS derived from solvent-free sample preparation

Trimpin

Räder

Müllen

2006

International Journal of Mass Spectrometry

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“…Earlier studies on protein and peptide analysis by MALDI‐MS showed that surfactants such as SDS, Triton‐X, or CHAPS interfered with MS analysis by reducing analyte ion signals and degrading mass resolution 6–9. However, in MALDI the extent of surfactant interference is not dependent only on the type of surfactant used.…”

mentioning

confidence: 99%

Effects of common surfactants on protein digestion and matrix‐assisted laser desorption/ionization mass spectrometric analysis of the digested peptides using two‐layer sample preparation

Zhang

2004

Rapid Comm Mass Spectrometry

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While surfactants are commonly used in preparing protein samples, their presence in a protein sample can potentially affect the enzymatic digestion process and the subsequent analysis of the resulting peptides by mass spectrometry. The extent of the tolerance of matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) to surfactant interference in peptide analysis is very much dependent on the matrix/sample preparation method. In this work the effects of four commonly used surfactants, namely n-octyl glucoside (OG), Triton X-100 (TX-100), 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) and sodium dodecyl sulfate (SDS), for biological sample preparation on trypsin digestion and MALDI-MS of the resulting digest are examined in detail within the context of using a two-layer method for MALDI matrix/sample preparation. Non-ionic and mild surfactants, such as OG, TX-100 or CHAPS, are found to have no significant effect on trypsin digestion with surfactant concentrations up to 1%. However, TX-100 and CHAPS interfere with the subsequent peptide analysis by MALDI-MS and should be removed prior to peptide analysis. OG is an MS-friendly surfactant and no effect is observed for MALDI peptide analysis. The effect of SDS on trypsin digestion in terms of the number of peptides generated and the overall protein sequence coverage by these peptides is found to be protein dependent. The use of SDS to solubilize hydrophobic membrane proteins, followed by trypsin digestion in the presence of 0.1% SDS, results in a peptide mixture that can be analyzed directly by MALDI-MS. These peptides are shown to provide better sequence coverage compared with those obtained without the use of SDS in the case of bacteriorhodopsin, a very hydrophobic transmembrane protein. This work illustrates that MALDI-MS with the two-layer sample preparation method can be used for direct analysis of protein digests with no or minimum sample cleanup after proteins are digested in a solution containing surfactants.

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“…Though surfactants have been used to improve the solubility of peptides and proteins in aqueous mediaof surfactants, especially anionic surfactants like SDS, compromises both the signal-to-noise ratio and the mass accuracy of MALDI-MS analysis [17][18][19][20]. These limitations are presumed to arise due to poor incorporation of the protein in the crystalline matrix because the protein interacts with the surfactant more than with the matrix molecules [21].…”

mentioning

confidence: 99%

Interactions between sodium dodecyl sulfate micelles and peptides during matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) of proteolytic digests

Tummala

Green‐Church

Limbach

2005

J. Am. Soc. Mass Spectrom.

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Although sodium dodecyl sulfate (SDS) is routinely used as a denaturing agent for proteins, its presence is highly detrimental on the analysis of peptides and proteins by mass spectrometry. It has been found, however, that when SDS is present in concentrations near to or above its critical micelle concentration (CMC), improvements in the matrix-assisted laser desorption/ ionization mass spectrometry (MALDI-MS) analysis of peptide mixtures or hydrophobic proteins are obtained. To elucidate possible explanations for such improvements, here we have undertaken a study examining the effect of SDS micelles on peptide mixtures. Fluorescently labeled peptides were used as probes to determine whether hydrophobic or hydrophilic peptides interact exclusively with SDS micelles. In addition, four globular proteins were digested with trypsin and then various amounts of SDS were added before MALDI mass spectrometry. To examine the role of mixture complexity on the mass spectral results, the tryptic digest of bovine serum albumin was also fractionated according to hydrophobicity before SDS treatment. Results from these experiments suggest that micelle-peptide interactions increase peptide-matrix cocrystallization irrespective of analyte hydrophobicity. As these studies were performed using the dried-droplet method of sample spotting, the presence of micelles is also hypothesized to reduce Marangoni effects during the crystallization process. (J Am Soc Mass Spectrom 2005, 16, 1438 -1446

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Approaches to the characterization of membrane channel proteins (porins) by UV MALDI-MS

Cited by 28 publications

References 32 publications

Investigations of theoretical principles for MALDI-MS derived from solvent-free sample preparation

Investigations of theoretical principles for MALDI-MS derived from solvent-free sample preparation

Effects of common surfactants on protein digestion and matrix‐assisted laser desorption/ionization mass spectrometric analysis of the digested peptides using two‐layer sample preparation

Interactions between sodium dodecyl sulfate micelles and peptides during matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) of proteolytic digests

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