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• 148757-94-2 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate 

Relevant academic research and scientific papers

Measurement of 15N enrichment of glutamine and urea cycle amino acids derivatized with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate using liquid chromatography-tandem quadrupole mass spectrometry

Abstract 6-Aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) is an amino acid-specific derivatizing reagent that has been used for sensitive amino acid quantification by liquid chromatography-tandem quadrupole mass spectrometry (LC-MS/MS). In this study, we aimed to evaluate the ability of this method to measure the isotopic enrichment of amino acids and to determine the positional 15N enrichment of urea cycle amino acids (i.e., arginine, ornithine, and citrulline) and glutamine. The distribution of the M and M + 1 isotopomers of each natural AQC-amino acid was nearly identical to the theoretical distribution. The standard deviation of the (M + 1)/M ratio for each amino acid in repeated measurements was approximately 0.1%, and the ratios were stable regardless of the injected amounts. Linearity in the measurements of 15N enrichment was confirmed by measuring a series of 15N-labeled arginine standards. The positional 15N enrichment of urea cycle amino acids and glutamine was estimated from the isotopic distribution of unique fragment ions generated at different collision energies. This method was able to identify their positional 15N enrichment in the plasma of rats fed 15N-labeled glutamine. These results suggest the utility of LC-MS/MS detection of AQC-amino acids for the measurement of isotopic enrichment in 15N-labeled amino acids and indicate that this method is useful for the study of nitrogen metabolism in living organisms.

Enhanced carboxypeptidase efficacies and differentiation of peptide epimers

Carboxypeptidases enzymatically cleave the peptide bond of C-terminal amino acids. In humans, it is involved in enzymatic synthesis and maturation of proteins and peptides. Carboxypeptidases A and Y have difficulty hydrolyzing the peptide bond of dipeptides and some other amino acid sequences. Early investigations into different N-blocking groups concluded that larger moieties increased substrate susceptibility to peptide bond hydrolysis with carboxypeptidases. This study conclusively demonstrates that 6-aminoquinoline-N-hydroxysuccimidyl carbamate (AQC) as an N-blocking group greatly enhances substrate hydrolysis with carboxypeptidase. AQC addition to the N-terminus of amino acids and peptides also improves chromatographic peak shapes and sensitivities via mass spectrometry detection. These enzymes have been used for amino acid sequence determination prior to the advent of modern proteomics. However, most modern proteomic methods assume that all peptides are comprised of L-amino acids and therefore cannot distinguish L-from D-amino acids within the peptide sequence. The majority of existing methods that allow for chiral differentiation either require synthetic standards or incur racemization in the process. This study highlights the resistance of D-amino acids within peptides to enzymatic hydrolysis by Carboxypeptidase Y. This stereoselectivity may be advantageous when screening for low abundance peptide stereoisomers.

Derivatize, Racemize, and Analyze - An Easy and Simple Procedure for Chiral Amino Acid Standard Preparation for Enantioselective Metabolomics

A simple, controllable, and reproducible stereoisomerization (racemization and epimerization) protocol for the preparation of scalemic α-amino acid mixtures from stereoisomerically pure standards was developed. Simply derivatize your amino acids with a racemization tag that incorporates a urea bond on the N-terminus of the target amino acid and incubate at elevated temperatures up to 95 °C for defined time periods until the targeted d-amino acid levels are obtained. The racemization tags investigated were 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC), aminophenyl-N-hydroxysuccinimidyl carbamate (AC), and 3-aminopyridyl-N-hydroxysuccinimidyl carbamate (APC). Employing this method, it was possible to create a ready-to-use, tailor-made chiral uniformly 13C and 15N labeled [U-13C15N]-amino acid standard with the desired d-amino acid percentage within minutes or hours without sample cleanup. A racemization time of 30 min at 95 °C will lead to a d-amino acid level of 1-5%, while 6 h at 95 °C provides 15-30% d-amino acids. Racemization occurs due to imine formation at the chiral carbon atom bound to the urea-linking group without decomposition of labile amino acids such as Asn, Gln, Trp, Cit, and theanine. For amino acids possessing two chiral centers such as dl-Ile or dl-Thr, only the epimerization of isomers with different stereochemistry at the second chiral center will produce all four possible isobaric enantiomers. All measurements were performed on the zwitterionic Chiralpak ZWIX(+) column using a dual hydro-organic flow gradient combined with HPLC-ESI-QTOF-MS analysis. This new racemization method solves the problem of (enantioselective) matrix effects and inaccurate results in LC-MS based enantioselective metabolomics and warrants full MS-compatibility.

Determination of isotopic labeling of proteins by precursor ion scanning liquid chromatography/tandem mass spectrometry of derivatized amino acids applied to nuclear magnetic resonance studies

RATIONALE A method has been developed for the quantitation of isotopic labeling of proteins using liquid chromatography/tandem mass spectrometry (LC/MS/MS) for the application of protein nuclear magnetic resonance (NMR) studies. NMR relies on specific isotopic nuclei, such as 13C and 15N, for detection and, therefore, isotopic labeling is an important sample preparation step prior to in-depth structural characterization of proteins. The goal of this study was to develop a robust quantitative assay for assessing isotopic labeling in proteins while retaining information on the extent of labeling for individual amino acids. METHODS Complete digestion of proteins by acid hydrolysis was followed by derivatization of free amino acids with 6-aminoquinolyl N-hydroxysuccinimidyl carbamate (AQC) forming derivatives having identical MS/MS fragmentation behavior. Precursor ion scanning on a hybrid quadrupole-linear ion trap platform was used for amino acid analysis and determining isotopic labeling of proteins. RESULTS Using a set of isotope-labeled amino acid standards mixed with their unlabeled counterparts, the method was validated for accurately measuring % isotopic contribution. We then applied the method for determining the 13C isotopic content of algal proteins during a feeding study using 13C6-glucose- or 13C-bicarbonate-supplemented culture media as well as the level of labeling in mussel byssal threads obtained after feeding with labeled algae. CONCLUSIONS This method is ideally suited for assessing the extent of protein labeling prior to NMR studies, where the isotopic labeling is a determining factor in the quality of resulting protein spectra, and can be applied to a multitude of different biological samples. Copyright