Tandem mass spectrometry performed on a pool of 18 oligopeptides shows that the product ion spectra of argentinated peptides, the [bn + OH + Ag]+ ions and the [yn - H + Ag]+ ions bearing identical sequences are virtually identical. These observations suggest strongly that these ions have identical structures in the gas phase. The structures of argentinated glycine, glycylglycine, and glycylglycylglycine were calculated using density functional theory (DFT) at the B3LYP/DZVP level of theory; they were independently confirmed using HF/LANL2DZ. For argentinated glycylglycylglycine, the most stable structure is one in which Ag+ is tetracoordinate and attached to the amino nitrogen and the three carbonyl oxygen atoms. Mechanisms are proposed for the fragmentation of this structure to the [b2 + OH + Ag]+ and the [Y2 - H + Ag]+ ions that are consistent with all experimental observations and known calculated structures and energetics. The structures of the [b2 - H + Ag]+ and the [a2 - H + Ag]+ ions of glycylglycylglycine were also calculated using DFT. These results confirm earlier suggestions that the [b2 - H + Ag]+ ion is an argentinated oxazolone and the [a2 - H + Ag]+ an argentinated immonium ion.
A strategy for semiautomatic sequencing of argentinated (silver-containing) oligopeptides has been developed. Sequencing is based on a search algorithm that identifies a triplet peak relationship in a product ion spectrum of the [M + Ag]+ ion of an oligopeptide. The ions that constitute a triplet are [bn + OH + Ag]+, [bn - H + Ag]+, and [a(n) - H + Ag]+, which are separated by 18 and 28 m/z units, respectively. The difference in the m/z values of adjacent triplets identifies the residue that is "cleaved". Observation of the [yn + H + Ag]+ ion containing the cleaved residue confirms the assignment. Sequencing of argentinated tryptic peptides may prove useful for automated proteome analysis via the sequence tag method.
The Multi-Attribute
Method (MAM) Consortium was initially formed
as a venue to harmonize best practices, share experiences, and generate
innovative methodologies to facilitate widespread integration of the
MAM platform, which is an emerging ultra-high-performance liquid chromatography–mass
spectrometry application. Successful implementation of MAM as a purity-indicating
assay requires new peak detection (NPD) of potential process- and/or
product-related impurities. The NPD interlaboratory study described
herein was carried out by the MAM Consortium to report on the industry-wide
performance of NPD using predigested samples of the NISTmAb Reference
Material 8671. Results from 28 participating laboratories show that
the NPD parameters being utilized across the industry are representative
of high-resolution MS performance capabilities. Certain elements of
NPD, including common sources of variability in the number of new
peaks detected, that are critical to the performance of the purity
function of MAM were identified in this study and are reported here
as a means to further refine the methodology and accelerate adoption
into manufacturer-specific protein therapeutic product life cycles.
Argentinated peptide ions are formed in abundance under matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) conditions in the presence of Ag+ ions. These argentinated peptide ions are fragmented facilely under MALDI-MS/MS conditions to yield [b(n) + OH + Ag]+, [b(n) - H + Ag]+ and [a(n) - H + Ag]+ ions that are indicative of the C-terminal sequence. These observations parallel those made earlier under electrospray MS conditions (Chu, I. K; Guo, X.; Lau, T.-C.; Siu, K W. M. Anal. Chem. 1999, 71, 2364-2372). A mixed protonated and argentinated tryptic peptide map was generated from 37 fmol of bovine serum albumin (BSA) using MALDI-MS. MALDI-MS/MS data from four argentinated peptides at a protein amount of 350 fmol unambiguously identified the protein as BSA. Sequence-tag analysis of two argentinated tryptic peptides was used to identify unambiguously myocyte enhancer factor 2A, which had been recombinantly expressed in a bacterial cell line.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.