Quantitative <sup>1</sup>H Nuclear Magnetic Resonance (qNMR) of Aromatic Amino Acids for Protein Quantification
Hydrolysis of protein samples into amino acids facilitates the use of NMR spectroscopy for protein and peptide quantification. Different conditions have been tested for quantifying aromatic amino acids and proteins. The pH-dependent signal shifts in the aromatic region of amino acid samples were exa...
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MDPI AG
2023-01-01
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| Series: | Methods and Protocols |
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| Online Access: | https://www.mdpi.com/2409-9279/6/1/11 |
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| author | Teodor Tchipilov Klas Meyer Michael G. Weller |
| author_facet | Teodor Tchipilov Klas Meyer Michael G. Weller |
| author_sort | Teodor Tchipilov |
| collection | DOAJ |
| description | Hydrolysis of protein samples into amino acids facilitates the use of NMR spectroscopy for protein and peptide quantification. Different conditions have been tested for quantifying aromatic amino acids and proteins. The pH-dependent signal shifts in the aromatic region of amino acid samples were examined. A pH of 12 was found to minimize signal overlap of the four aromatic amino acids. Several aromatic compounds, such as terephthalic acid, sulfoisophthalic acid, and benzene tricarboxylic acid, were applied as internal standards. The quantification of amino acids from an amino acid standard was performed. Using the first two suggested internal standards, recovery was ~97% for histidine, phenylalanine, and tyrosine at a concentration of approximately 1 mM in solution. Acidic hydrolysis of a certified reference material (CRM) of bovine serum albumin (BSA) and subsequent quantification of Phe and Tyr yielded recoveries of 98% ± 2% and 88% ± 4%, respectively, at a protein concentration of 16 g/L or 250 µM. |
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| language | English |
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| spelling | doaj.art-2901d983935e458d9df7ca13ab0247a82023-11-16T22:26:18ZengMDPI AGMethods and Protocols2409-92792023-01-01611110.3390/mps6010011Quantitative <sup>1</sup>H Nuclear Magnetic Resonance (qNMR) of Aromatic Amino Acids for Protein QuantificationTeodor Tchipilov0Klas Meyer1Michael G. Weller2Federal Institute for Materials Research and Testing (BAM), Division 1.5 Protein Analysis, Richard Willstätter-Strasse 11, 12489 Berlin, GermanyFederal Institute for Materials Research and Testing (BAM), Division 1.4 Process Analytical Technology, Richard Willstätter-Strasse 11, 12489 Berlin, GermanyFederal Institute for Materials Research and Testing (BAM), Division 1.5 Protein Analysis, Richard Willstätter-Strasse 11, 12489 Berlin, GermanyHydrolysis of protein samples into amino acids facilitates the use of NMR spectroscopy for protein and peptide quantification. Different conditions have been tested for quantifying aromatic amino acids and proteins. The pH-dependent signal shifts in the aromatic region of amino acid samples were examined. A pH of 12 was found to minimize signal overlap of the four aromatic amino acids. Several aromatic compounds, such as terephthalic acid, sulfoisophthalic acid, and benzene tricarboxylic acid, were applied as internal standards. The quantification of amino acids from an amino acid standard was performed. Using the first two suggested internal standards, recovery was ~97% for histidine, phenylalanine, and tyrosine at a concentration of approximately 1 mM in solution. Acidic hydrolysis of a certified reference material (CRM) of bovine serum albumin (BSA) and subsequent quantification of Phe and Tyr yielded recoveries of 98% ± 2% and 88% ± 4%, respectively, at a protein concentration of 16 g/L or 250 µM.https://www.mdpi.com/2409-9279/6/1/11amino acid analysisAAAprotein hydrolysismetrologytraceabilityreference materials |
| spellingShingle | Teodor Tchipilov Klas Meyer Michael G. Weller Quantitative <sup>1</sup>H Nuclear Magnetic Resonance (qNMR) of Aromatic Amino Acids for Protein Quantification Methods and Protocols amino acid analysis AAA protein hydrolysis metrology traceability reference materials |
| title | Quantitative <sup>1</sup>H Nuclear Magnetic Resonance (qNMR) of Aromatic Amino Acids for Protein Quantification |
| title_full | Quantitative <sup>1</sup>H Nuclear Magnetic Resonance (qNMR) of Aromatic Amino Acids for Protein Quantification |
| title_fullStr | Quantitative <sup>1</sup>H Nuclear Magnetic Resonance (qNMR) of Aromatic Amino Acids for Protein Quantification |
| title_full_unstemmed | Quantitative <sup>1</sup>H Nuclear Magnetic Resonance (qNMR) of Aromatic Amino Acids for Protein Quantification |
| title_short | Quantitative <sup>1</sup>H Nuclear Magnetic Resonance (qNMR) of Aromatic Amino Acids for Protein Quantification |
| title_sort | quantitative sup 1 sup h nuclear magnetic resonance qnmr of aromatic amino acids for protein quantification |
| topic | amino acid analysis AAA protein hydrolysis metrology traceability reference materials |
| url | https://www.mdpi.com/2409-9279/6/1/11 |
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