Electron Transfer Dissociation of Peptides Generated by Microwave D-Cleavage Digestion of Proteins

Hauser, N.; Han, Hongling; McLuckey, Scott A.; Basile, Franco

doi:10.1021/pr700671z

Cited by 32 publications

(27 citation statements)

References 21 publications

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“…By using LC-ESI-LTQ-Orbitrap and acetic acid digestion, Swatkoski et al showed that database search with CID MS/MS (for small peptide) was successful with some modification on the MAS-COT enzyme rules [6]. For larger and highly charged Aspcleavage peptides, ETD (and ECD) was found by Hauser et al to be more suitable than CID to yield larger sequence coverage [37].…”

Section: Resultsmentioning

confidence: 99%

Rapid Online Non-Enzymatic Protein Digestion Analysis with High Pressure Superheated ESI-MS

Chen

Kinoshita

Noda

et al. 2015

J. Am. Soc. Mass Spectrom.

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Abstract. Recently, we reported a new ESI ion source that could electrospray the super-heated aqueous solution with liquid temperature much higher than the normal boiling point (J. Am. Soc. Mass Spectrom. 25, 1862-1869). The boiling of liquid was prevented by pressurizing the ion source to a pressure greater than atmospheric pressure. The maximum operating pressure in our previous prototype was 11atm, and the highest achievable temperature was 180°C. In this paper, a more compact prototype that can operate up to 27atm and 250°C liquid temperatures is constructed, and reproducible MS acquisition can be extended to electrospray temperatures that have never before been tested. Here, we apply this super-heated ESI source to the rapid online protein digestion MS. The sample solution is rapidly heated when flowing through a heated ESI capillary, and the digestion products are ionized by ESI in situ when the solution emerges from the tip of the heated capillary. With weak acid such as formic acid as solution, the thermally accelerated digestion (acid hydrolysis) has the selective cleavage at the aspartate (Asp, D) residue sites. The residence time of liquid within the active heating region is about 20s. The online operation eliminates the need to transfer the sample from the digestion reactor, and the output of the digestive reaction can be monitored and manipulated by the solution flow rate and heater temperature in a near real-time basis.

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Section: Resultsmentioning

confidence: 99%

Rapid Online Non-Enzymatic Protein Digestion Analysis with High Pressure Superheated ESI-MS

Chen

Kinoshita

Noda

et al. 2015

J. Am. Soc. Mass Spectrom.

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“…As reported by Hauser et al microwave digestion occurs very fast (,6 min) and subsequent ETD analysis results in sufficient fragment ions for confident peptide identification. The performance and rapidness of the method are ideal for an ETD application in high-throughput proteomics [43].…”

Section: Other Proteasesmentioning

confidence: 99%

Application of electron transfer dissociation (ETD) for the analysis of posttranslational modifications

Wiesner¹,

Premsler²,

2008

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Despite major advantages in the field of proteomics, the analysis of PTMs still poses a major challenge; thus far, preventing insights into the role and regulation of protein networks. Additionally, top-down sequencing of proteins is another powerful approach to reveal comprehensive information for biological function. A commonly used fragmentation technique in MS-based peptide sequencing is CID. As CID often fails in PTM-analysis and performs best on doubly-charged, short and middle-sized peptides, confident peptide identification may be hampered. A newly developed fragmentation technique, namely electron transfer dissociation (ETD), supports both, PTM- and top-down analysis, and generally results in more confident identification of long, highly charged or modified peptides. The following review presents the theoretical background of ETD and its technical implementation in mass analyzers. Furthermore, current improvements of ETD and approaches for the PTM-analysis and top-down sequencing are introduced. Alternating both fragmentation techniques, ETD and CID, increases the amount of information derived from peptide fragmentation, thereby enhancing both, peptide sequence coverage and the confidence of peptide and protein identification.

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“…As a result, these CID mass spectra are not as informative in terms of amino acid sequence because the localization of charges on basic groups along the backbone of the peptide does not allow for sufficiently random fragmentation. In fact, the high charge state peptides produced by the microwave D-cleavage method have been shown to be better suited for Electron Transfer Dissociation (ETD) than for CID, the former technique favoring ions with charge states higher than +2, and thus providing a high percentage of sequence coverage32. Conversely, direct comparison of CID tandem mass spectra of peptides generated by microwave D-cleavege and trypsin digestion showed improved peptide sequence coverage for the tryptic peptide over the microwave-generated peptide32.…”

Section: Introductionmentioning

confidence: 99%

“…In fact, the high charge state peptides produced by the microwave D-cleavage method have been shown to be better suited for Electron Transfer Dissociation (ETD) than for CID, the former technique favoring ions with charge states higher than +2, and thus providing a high percentage of sequence coverage32. Conversely, direct comparison of CID tandem mass spectra of peptides generated by microwave D-cleavege and trypsin digestion showed improved peptide sequence coverage for the tryptic peptide over the microwave-generated peptide32. This behavior is expected to be similar for the echem WY-cleavage derived peptides since tryptophan and tyrosine have a combined frequency in protein sequences of 3.99 %31.…”

Section: Introductionmentioning

confidence: 99%

Rapid Online Nonenzymatic Protein Digestion Combining Microwave Heating Acid Hydrolysis and Electrochemical Oxidation

Basile

Hauser

2010

Anal. Chem.

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We report an online non-enzymatic method for site-specific digestion of proteins to yield peptides that are well suited for collision induced dissociation (CID) tandem mass spectrometry (MS/MS). The method combines online microwave heating acid hydrolysis at aspartic acid and online electrochemical oxidation at tryptophan and tyrosine. The combined microwave/electrochemical (microwave/echem) digestion is reproducible and produces peptides with an average sequence length of 10 amino acids. This peptide length is similar to the average peptide length of 9 amino acids obtained by digestion of proteins with the enzyme trypsin. As a result, the peptides produced by this novel non-enzymatic digestion method, when analyzed by ESI-MS, produce protonated molecules with mostly +1 and +2 charge states. The combination of these two non-enzymatic methods overcomes shortcomings with each individual method in that: i) peptides generated by the microwave-hydrolysis method have an average amino acid length of 16 amino acids, and ii) the inability of the electrochemical-cleavage method to reproducibly digest proteins with molecular masses above 4 kDa. Preliminary results are presented on the application and utility of this rapid online digestion (total of 6 min digestion time) on a series of standard peptides and proteins as well as an E. coli protein extract.

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Electron Transfer Dissociation of Peptides Generated by Microwave D-Cleavage Digestion of Proteins

Cited by 32 publications

References 21 publications

Rapid Online Non-Enzymatic Protein Digestion Analysis with High Pressure Superheated ESI-MS

Rapid Online Non-Enzymatic Protein Digestion Analysis with High Pressure Superheated ESI-MS

Application of electron transfer dissociation (ETD) for the analysis of posttranslational modifications

Rapid Online Nonenzymatic Protein Digestion Combining Microwave Heating Acid Hydrolysis and Electrochemical Oxidation

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