Materials and methods for detection and quantification of beta-n-methylamino-l-alanine
AssigneeNorth Carolina State University
InventorsDavid C. Muddiman, Philip L. Loziuk, Joshua G. Pierce, Gavin Williams, Yasamin Moazami
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Abstract
Provided herein are isotopically labeled reagents, including isotopically labeled small molecules and peptides, that can be used to detect and/or quantify β-N-methylamino-L-alanine (BMAA) in a sample. The reagents can be used as stable isotope labeled standards in analytical methods, including in conjunction with mass spectrometry, to detect and/or quantify BMAA in a sample, such as a protein sample from a subject.
Claims36
A composition comprising an isotopically labeled compound defined by the formula below
- wherein R represents hydrogen or an amine protecting group;
- and
- wherein at least two of aC, bC, cC, dC, eN, fN, gO, and hO are isotopically labeled with a stable isotope.
A composition comprising a polypeptide, wherein the polypeptide includes one or more isotopically labeled β-N-methylamino-L-alanine (BMAA) residues, wherein each of the one or more isotopically labeled BMAA residues is isotopically labeled with one or more stable isotopes.
A method for quantifying the amount of β-N-methylamino-L-alanine (BMAA) in a sample comprising:
- (i) analyzing a test sample by mass spectrometry;
- (ii) spiking the test sample with a defined amount of an isotopically labeled compound defined by the formula below
- wherein R represents hydrogen or an amine protecting group, and at least two of aC, bC, cC, dC, eN, fN, gO, and hO are isotopically labeled with a stable isotope;
- to provide a BMAA-spiked sample;
- (iii) analyzing the BMAA-spiked sample by mass spectrometry;
- and
- (iv) determining the amount of BMAA in the test sample by isotope dilution analysis.
A method for incorporating an isotopically-labeled β-N-methylamino-L-alanine (BMAA) residue into a protein or peptide, the method comprising:
- providing a protein or peptide that includes a phosphoserine residue;
- reacting the protein or peptide to convert the phosphoserine residue to an α,β-unsaturated amino acid residue;
- and
- reacting the α,β-unsaturated amino acid residue with methylamine to provide a BMAA residue, wherein the methylamine is isotopically enriched with one or more stable isotopes.
Dependencies are inferred from the claim text.