کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
1220486 1494619 2015 8 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
The stereoselective separation of serine containing peptides by zwitterionic ion exchanger type chiral stationary phases and the study of serine racemization mechanisms by isotope exchange and tandem mass spectrometry
موضوعات مرتبط
مهندسی و علوم پایه شیمی شیمی آنالیزی یا شیمی تجزیه
پیش نمایش صفحه اول مقاله
The stereoselective separation of serine containing peptides by zwitterionic ion exchanger type chiral stationary phases and the study of serine racemization mechanisms by isotope exchange and tandem mass spectrometry
چکیده انگلیسی


• Direct enantioseparation of di-, tri- and tetrapeptides by novel ZWIX CSPs.
• Baseline HPLC separation of d-AA containing peptides and detection by MS2.
• Ser racemization mechanisms studied by HRMS2 measurements and isotope exchange.
• Ser in peptides racemizes via Cα proton exchange or via β-elimination of OH.

The occurrence of d-amino acids (d-AAs) in higher-developed organisms in their free form, and within peptides and proteins, has been investigated with an increasing number of studies. Often the inversion of the stereochemical configuration of an individual amino acid drastically changes its biological activity. Alongside Asn and Asp, Ser is most prone to racemization within peptides. Specific enzymes catalyzing d-Ser generation and breakdown have been described. Hence, the applicability of enantioselective ZWIX(+)® and ZWIX(−)® chiral stationary phases (CSPs) to peptide separations was assessed and a set of 14 pairs of diastereomeric and enantiomeric Ser and Thr containing di-, tri- and tetra-peptides was chromatographically separated without prior hydrolysis to the individual amino acids. To a certain extent, RP chromatography also enabled the separation of peptide diastereomers. The ZWIX CSPs delivered chromatographic selectivities between 1.04 and 7.23, allowing a change of elution order by switching between the ZWIX(+) and the ZWIX(−) CSP. Coupling these highly selective chromatographic columns with an LTQ-Orbitrap XL™ mass spectrometer and performing high resolution MS2 measurements enabled us to investigate mechanistic aspects of chemically induced racemization of Ser embedded in short peptides. As reaction medium an alkaline aqueous solution (pH 12.3) was selected. Proton/deuterium exchange experiments provided evidence of a fast Cα proton exchange with simultaneous racemization. Additionally, 18O/16O exchange allowed the identification of an alternative, and somewhat retarded racemization via a reversible β-elimination and reintroduction of water at the hydroxymethyl side chain of Ser. This involved the intermediate generation of the prochiral didehydro alanine unit.

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ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Journal of Pharmaceutical and Biomedical Analysis - Volume 116, 10 December 2015, Pages 123–130
نویسندگان
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