Kinetic Characterization of Lysine-Specific Metalloendopeptidases from Grifola frondosa and Pleurotus ostreatus Fruiting Bodies

Nonaka, Takashi; Hashimoto, Yohichi; Takio, Koji

doi:10.1093/oxfordjournals.jbchem.a022074

Cited by 38 publications

(45 citation statements)

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“…Taking into account that most of the proteomic studies are currently carried out according to shotgun strategies involving the collision-induced dissociations in ESI-MS/MS of doublyand triply charged precursor ions [58], the described exclusion phenomenon mainly concerns sequencing methodologies based on MALDI-TOF/TOF analyses. This atypical 50 100 150 200 250 300 350 400 450 500 550 600 650 700 750 800 850 900 950 1000 1050 fragmentation might be deleterious for efficient correct sequence assignment, especially when proteolysis is carried out with others enzymes than trypsin such as Lys-N [28], Asp-N [59], and Glu-C [60] proteases, which produce peptides bearing arginine and histidine anywhere within the proteolytic sequences.…”

Section: Influence Of the Charge State Of The Precursor Ion On C-termmentioning

confidence: 99%

“…As depicted in Figure 1, among the peptide free acids, some were designed to mimic protein digest sequences such as tryptic chains having either arginine (R) or lysine (K) at their C-terminus [27], as well as Lys-N proteolytic peptides that all possess N-terminal lysine [28,29]. Peptides illustrating trypsin miscleavages with either R or K within the sequence were also exemplified.…”

Section: Introductionmentioning

confidence: 99%

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Occurrence of C-Terminal Residue Exclusion in Peptide Fragmentation by ESI and MALDI Tandem Mass Spectrometry

Dupré

Cantel

Martinez

et al. 2011

J. Am. Soc. Mass Spectrom.

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By screening a data set of 392 synthetic peptides MS/MS spectra, we found that a known C-terminal rearrangement was unexpectedly frequently occurring from monoprotonated molecular ions in both ESI and MALDI tandem mass spectrometry upon low and high energy collision activated dissociations with QqTOF and TOF/TOF mass analyzer configuration, respectively. Any residue localized at the C-terminal carboxylic acid end, even a basic one, was lost, provided that a basic amino acid such arginine and to a lesser extent histidine and lysine was present in the sequence leading to a fragment ion, usually depicted as (b n-1 +H 2 O) ion, corresponding to a shortened nonscrambled peptide chain. Far from being an epiphenomenon, such a residue exclusion from the peptide chain C-terminal extremity gave a fragment ion that was the base peak of the MS/MS spectrum in certain cases. Within the frame of the mobile proton model, the ionizing proton being sequestered onto the basic amino acid side chain, it is known that the charge directed fragmentation mechanism involved the C-terminal carboxylic acid function forming an anhydride intermediate structure.The same mechanism was also demonstrated from cationized peptides. To confirm such assessment, we have prepared some of the peptides that displayed such C-terminal residue exclusion as a C-terminal backbone amide. As expected in this peptide amide series, the production of truncated chains was completely suppressed. Besides, multiply charged molecular ions of all peptides recorded in ESI mass spectrometry did not undergo such fragmentation validating that any mobile ionizing proton will prevent such a competitive C-terminal backbone rearrangement. Among all well-known nondirect sequence fragment ions issued from non specific loss of neutral molecules (mainly H 2 O and NH 3 ) and multiple backbone amide ruptures (b-type internal ions), the described Cterminal residue exclusion is highly identifiable giving raise to a single fragment ion in the high mass range of the MS/MS spectra. The mass difference between this signal and the protonated molecular ion corresponds to the mass of the C-terminal residue. It allowed a straightforward identification of the amino acid positioned at this extremity. It must be emphasized that a neutral residue loss can be misattributed to the formation of a y m-1 ion, i.e., to the loss of the N-terminal residue following the a 1 -y m-1 fragmentation channel. Extreme caution must be adopted when reading the direct sequence ion on the positive ion MS/MS spectra of singly charged peptides not to mix up the attribution of the N-and C-terminal amino acids. Although such peculiar fragmentation behavior is of obvious interest for de novo peptide sequencing, it can also be exploited in proteomics, especially for studies involving digestion protocols carried out with proteolytic enzymes other than trypsin (Lys-N, Glu-C, and Asp-N) that produce arginine-containing peptides.

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Section: Influence Of the Charge State Of The Precursor Ion On C-termmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Occurrence of C-Terminal Residue Exclusion in Peptide Fragmentation by ESI and MALDI Tandem Mass Spectrometry

Dupré

Cantel

Martinez

et al. 2011

J. Am. Soc. Mass Spectrom.

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“…Alternative to the above described chemical approaches, we recently introduced a biochemical approach to manipulate the basicity of the peptide termini, using the metalloendopeptidase Lys-N [13][14][15]. This enzyme has cleavage specificity N-terminal of lysine residues [16], creating peptides with a strong basicity on the N-terminus caused by the presence of the N-terminal lysine. Fragmentation of singly charged Lys-N peptides that contain a lysine as the only basic residue generates spectra dominated by N-terminal fragment ions.…”

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confidence: 99%

“…An easy way of specific C-terminal labeling has been accomplished by proteolytic 18 O incorporation with the protease trypsin. Fragmentation of the 16 O and 18 O labeled peptides, either individually and/or simultaneously, enables the identification of fragments from the 16 O/ 18 O C-terminus and has been used for de novo sequencing [21][22][23][24]. Most of the N-terminal labeling strategies target primary amines, as several specific derivatization procedures are known [10,11,25].…”

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confidence: 99%

Dimethyl isotope labeling assisted de novo peptide sequencing

Hennrich

Mohammed

Altelaar

et al. 2010

J. Am. Soc. Mass Spectrom.

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Here, we explore a de novo sequencing strategy in which we combine Lys-N protein digestion with differential isotopic dimethyl labeling to facilitate the (de novo) identification of multiply charged peptides in ESI-MS, both under CID and ETD conditions. For a large fraction of the Lys-N generated peptides, all primary amines are present at the N-terminal lysine, enabling specific labeling of the N-terminus. Differential derivatization of only the peptide N-terminus in combination with the simultaneous fragmentation of the corresponding isotopologues allows the straightforward distinction of N-terminal fragments from C-terminal and internal fragments. Furthermore, also singly and multiply charged N-terminal fragments can easily be distinguished due to the mass differences of the isotope labeled fragment pairs. As a proof of concept, we applied this approach to proteins isolated from an avocado fruit, and were able to partially de novo sequence and correctly align, with green plant homologues, a previously uncharacterized avocado ascorbate peroxidase. (J Am Soc Mass Spectrom 2010, 21, 1957-1965

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“…The proteolytic activity of a number of aspzincins has been characterised and, in certain bacterial and fungal pathogenic systems, aspzincin activity is highly toxic to mice and fish (Arnadottir et al 2009;Yamada et al 2012). Characterised aspzincins include lysine-specific metalloendopeptidases from the fungi Grifola frondosa and Pleurotus ostreatus (Nonaka et al 1998), deuterolysin from fungus Aspergillus oryzae (Fushimi et al 1999), the aspzincin AsaP1 in bacterium Aeromonas salmonicida subsp. achromogenes (Arnadottir et al 2009), and penicillolysin from Penicillium citrinum (Doi et al 2004).…”

Section: Introductionmentioning

confidence: 99%

Horizontal gene transfer in the sponge Amphimedon queenslandica

Higgie¹

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Horizontal gene transfer (HGT) is the nonsexual transfer of genetic sequence across species boundaries.Historically, HGT has been assumed largely irrelevant to animal evolution, though widely recognised as an important evolutionary force in bacteria. From the recent boom in whole genome sequencing, many cases have emerged strongly supporting the occurrence of HGT in a wide range of animals.However, the extent, nature and mechanisms of HGT in animals remain poorly understood. Here, I explore these uncertainties using 576 HGTs previously reported in the genome of the demosponge Amphimedon queenslandica.The HGTs derive from bacterial, plant and fungal sources, contain a broad range of domain types, and many are differentially expressed throughout development. Some domains are highly enriched; phylogenetic analyses of the two largest groups, the Aspzincin_M35 and the PNP_UDP_1 domain groups, suggest that each results from one or few transfer events followed by post-transfer duplication.Their differential expression through development, and the conservation of domains and duplicates, together suggest that many of the HGT-derived genes are functioning in A. queenslandica. The largest group consists of aspzincins, a metallopeptidase found in bacteria and fungi, but not typically in animals.I detected aspzincins in representatives of all four of the sponge classes, suggesting that the original sponge aspzincin was transferred after sponges diverged from their last common ancestor with the Eumetazoa, but before the contemporary sponge classes emerged. In A. queenslandica, the aspzincins may have been co-opted for multiple functions, since 54 of the 90 fit into one of four ontogenetic expression profiles, each of which is putatively co-expressed with different suites of native genes.Based on secretion signals and the conservation of key catalytic residues, I propose that proteolytic activity is maintained in at least one of the aspzincin roles in A. queenslandica.Mobile elements are capable of genomic excision, movement and integration, they enable HGT in bacteria, and some animal HGTs are genomically close to eukaryotic transposable elements (TEs); as such, mobile elements are speculated as possible players in the mechanisms of interkingdom and interdomain HGT to animals. In A. queenslandica, the overall repeats densities around predicted Together, these results offer novel insight on HGT in A. queenslandica and demonstrate that HGT has a varied nature in this animal with an extensive impact, partially due to post-transfer duplications. Further, this work offers insights on possible mechanisms that led to the HGT derived genes of this sponge. iv Declaration by authorThis thesis is composed of my original work, and contains no material previously published or written by another person except where due reference has been made in the text. I have clearly stated the contribution by others to jointly-authored works that I have included in my thesis.

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Kinetic Characterization of Lysine-Specific Metalloendopeptidases from Grifola frondosa and Pleurotus ostreatus Fruiting Bodies

Cited by 38 publications

References 0 publications

Occurrence of C-Terminal Residue Exclusion in Peptide Fragmentation by ESI and MALDI Tandem Mass Spectrometry

Occurrence of C-Terminal Residue Exclusion in Peptide Fragmentation by ESI and MALDI Tandem Mass Spectrometry

Dimethyl isotope labeling assisted de novo peptide sequencing

Horizontal gene transfer in the sponge Amphimedon queenslandica

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