Characterization of intermolecular ?-sheet peptides by mass spectrometry and hydrogen isotope exchange

Kraus, Mario; Janek, Katharina; Bienert, Michael; Krause, Eberhard

doi:10.1002/1097-0231(20000715)14:13<1094::aid-rcm994>3.0.co;2-5

Cited by 28 publications

(34 citation statements)

References 38 publications

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“…Studies with model peptides show that -sheet formation and increased length of the -strand motif increase the tendency to aggregate (29). Our data support these results insofar as the dodecapeptide KFFEAAAKKFFE forms more abundant and larger fibrils than the KFFE tetrapeptide.…”

Section: Discussionsupporting

confidence: 83%

Folding into a β-Hairpin Can Prevent Amyloid Fibril Formation

et al. 2004

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The tetrapeptide KFFE is one of the shortest amyloid fibril-forming peptides described. Herein, we have investigated how the structural environment of this motif affects polymerization. Using a turn motif (YNGK) or a less rigid sequence (AAAK) to fuse two KFFE tetrapeptides, we show by several biophysical methods that the amyloidogenic properties are strongly dependent on the structural environment. The dodecapeptide KFFEAAAKKFFE forms abundant thick fibril bundles. Freshly dissolved KFFEAAAKKFFE is monomeric and shows mainly disordered secondary structure, as evidenced by circular dichroism, NMR spectroscopy, hydrogen/deuterium exchange measurements, and molecular modeling studies. In sharp contrast, the dodecapeptide KFFEYNGKKFFE does not form fibrils but folds into a stable beta-hairpin. This structure can oligomerize into a stable 12-mer and multiples thereof, as shown by size exclusion chromatography, sedimentation analysis, and electrospray mass spectrometry. These data indicate that the structural context in which a potential fibril forming sequence is present can prevent fibril formation by favoring self-limiting oligomerization over polymerization.

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Section: Discussionsupporting

confidence: 83%

Folding into a β-Hairpin Can Prevent Amyloid Fibril Formation

et al. 2004

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“…As PSD makes a considerable contribution to the abundance of fragment ions in our MS/MS experiments, it is evident that complete scrambling has occurred in the metastable precursors that undergo PSD. In this context, we note that PSD has been used previously in the attempt to localize the sites of incorporation of deuterium in solution (19,20). In these studies, it was implicitly assumed that the level of scrambling was negligible, but no experimental evidence was presented to support this assumption.…”

Section: Discussionmentioning

confidence: 99%

“…For a first hand view, gas-phase fragmentation in the mass spectrometer may appear to be the logical choice for an auxiliary method providing the desired site-specific informa-tion. Accordingly it has been widely adopted in the attempt to identify specific sites that have become deuterated in solution 1 H/ 2 H exchange experiments (7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21). It is, however, mandatory for this approach that the level of intramolecular hydrogen ( 1 H/ 2 H) migration upon ion activation is negligible.…”

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confidence: 99%

Collisional Activation by MALDI Tandem Time-of-flight Mass Spectrometry Induces Intramolecular Migration of Amide Hydrogens in Protonated Peptides

Jørgensen

Bache

Roepstorff

et al. 2005

Molecular & Cellular Proteomics

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Mass spectrometry-based proteomics is currently one of the key technologies for the identification of protein interaction networks in the cell (1-3). This technology has provided a wealth of information unraveling the complexity of these biological processes at the protein level. A detailed understanding of the function of these protein networks at the molecular level requires, however, characterization of the dynamics and structural properties of the interacting proteins. Structural analyses by NMR spectroscopy or x-ray crystallography require relatively large amounts of pure proteins (typically milligram quantities), and many proteins are furthermore not inherently amenable to analysis by these traditional techniques (e.g. modular proteins or proteins with heterogeneous glycosylation patterns are often reluctant to yield well diffracting crystals).An alternative approach for the characterization of protein complexes without these limitations is provided by carefully monitoring by mass spectrometry changes in the rates of amide hydrogen ( 1 H/ 2 H) exchange upon complex formation (4, 5). In a typical experimental set-up for investigation of conformational properties of a protein complex, the unligated proteins as well as the preformed protein complex are incubated separately in deuterated buffer under physiological conditions. Global deuterium incorporation kinetics are subsequently established by monitoring deuterium contents in these protein samples withdrawn at appropriate intervals with their amide hydrogen isotopic exchange quenched by acidification and cooling. Information about local deuterium incorporation is subsequently collected after pepsin digestion followed by reversed-phase LC-MS. The chromatography is carried out with cold protiated solvents allowing effective back-exchange at all exchangeable sites with protium ( 1 H) with the sole exception of the amide groups forming the polypeptide chain. The actual number of individual amide hydrogens that can be resolved by this method, however, is limited by the number and sizes of peptides generated by pepsin. Overlapping sequence coverage is often provided by the broad specificity of pepsin, and although this promiscuity in substrate recognition increases the resolution in the local assignment of deuterium incorporation, single residue information can generally be obtained for only a few positions (6). For a first hand view, gas-phase fragmentation in the mass spectrometer may appear to be the logical choice for an auxiliary method providing the desired site-specific informaFrom the ‡Department

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“…The time-resolved HX experiments further support the existence of secondary structures for neoendorphins in the presence of TFE. In water, all of the labile hydrogens were exchanged within 30 s. This type of fast HX behavior is a common phenomenon for several other peptides [28][29][30][31][32][33][34][35]. The HX rates were relatively slower in 50% and 80% solvent systems.…”

Section: Discussionmentioning

confidence: 99%

“…The information about the number of exchanged hydrogens from the MS/MS spectra is derived from the m/z values of b and y ions by using the following expressions [33]. :…”

Section: Calculations Of Deuterium Incorporation In the Peptide Molecmentioning

confidence: 99%

Trifluoroethanol-induced conformational changes in α- and β-neoendorphins monitored using hydrogen/deuterium exchange mass spectrometry and circular dichroism spectroscopy

Kosanam

Dass

2011

International Journal of Mass Spectrometry

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Characterization of intermolecular ?-sheet peptides by mass spectrometry and hydrogen isotope exchange

Cited by 28 publications

References 38 publications

Folding into a β-Hairpin Can Prevent Amyloid Fibril Formation

Folding into a β-Hairpin Can Prevent Amyloid Fibril Formation

Collisional Activation by MALDI Tandem Time-of-flight Mass Spectrometry Induces Intramolecular Migration of Amide Hydrogens in Protonated Peptides

Trifluoroethanol-induced conformational changes in α- and β-neoendorphins monitored using hydrogen/deuterium exchange mass spectrometry and circular dichroism spectroscopy

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