Pyrylium‐based dye and charge tagging in proteomics

Abstract

The pyrylium group is a selective reagent for ε-amino groups in proteins. In particular, for fluorescence labeling, a number of advantages over traditional N-hydroxysuccinimidyl ester chemistry were recognized such as the rapid prestaining procedure. Here, we have investigated the labeling reaction for the fluorogenic pyrylium dye Py-1 using liquid chromatography coupled to MS with the aim of determining its specificity and possible side products. Peptides containing no, one, and two lysine residue and a choice of no or one cysteine residue were labeled with Py-1 at yields > 30%. Gas phase fragmentation proved both labeling of lysine residues as well as that of the N-terminus also in peptides that contained a lysine residue. Evidence for cysteine labeling was not found, but several other products were detected such as the results of rearrangements with adjacent acidic amino acids. Apart from the use as a fluorogenic label, Py-1 recommends itself for N-terminal charge tagging as alternative to the commonly used quaternary ammonium salts. Predominantly a- and b-type ion series were observed for N-terminally labeled peptides. Further applications include chromophore tagging since the labeled product is not only fluorescent but also colored red.

Upconversion NIR-II fluorophores for mitochondria-targeted cancer imaging and photothermal therapy

“…As useful and versatile heterocyclic luminescent compounds with a positive charge, pyrylium salts are widely utilized as photosensitizers, Q-switchers, and NIR dyes for proteins and oligosaccharide labeling 42,43 . Thiopyrylium is analogous to the pyrylium cation with the oxygen atom replaced by a sulfur atom.…”

Ultrasensitive Detection and Separation of Pancreatic Cancer Biomarker CA 19-9 Using a Multiphoton Laser Wave-Mixing Detector Interfaced to Capillary Electrophoresis

“…Chromeo P503 also degrades quickly due to ringopening in a basic solution, and hence, excess-dye optical absorption is negligible at 532. 35,37 Figure 7 shows UV−visible absorption spectra of Chromeo P503-conjugated CA 19-9 using various Chromeo P503 to CA 19-9 molar ratios and the corresponding amounts of Chromeo P503 without CA 19-9 dissolved in the same buffer. The net absorbance at 532 nm (absorbance of Chromeo P503conjugated CA 19-9 minus absorbance of degraded Chromeo P503) increases steadily when the molar ratio is increased from 1:1 to 5:1.…”

“…This study also uses Chromeo Py dyes, which are pyrylium dyes with heterocyclic aromatic rings with a bonded chromophore . Previous works showed pyrylium dyes converted to stable pyridium salts by reacting with the primary amine groups of the protein. , Most importantly, one can skip the dialysis step.…”

Enlightening the Path to Protein Engineering: Chemoselective Turn-On Probes for High-Throughput Screening of Enzymatic Activity

“…Although Py-1 31 is the most widely used derivative, − many other spectral bands are addressable by modification of the aromatic core as exemplified by the benzothiazole derivative 33 (Figure ). The reactivity of those salts could furthermore be fine-tuned by modifying the substituents of the pyrylium moiety .…”