Glycerol affects the quantification of aspartate and glutamate in acid-hydrolyzed proteins.
Glycerol is widely used in protein isolation pathways to improve folding and solubility of the proteins of interest. Amino acid composition analysis of protein samples hydrolyzed in the presence of glycerol resulted in underestimation of aspartate and glutamate, when compared to hydrolysis in the absence of glycerol. Quantification of free asparagine, aspartic acid, glutamine and glutamic acid hydrolyzed with hydrochloric acid or methanesulfonic acid in the presence of glycerol resulted in poor recoveries of aspartate and glutamate (between 6 and 66%). Gas chromatography-mass spectrometry analysis of the hydrolyzates revealed, as expected, the presence of esterification products. The esters were formed between the primary and secondary hydroxyl groups of the glycerol and both carboxyl groups of the amino acids. Protein samples intended for compositional analysis should be free of glycerol.
Juranville
J F
JF
F. Hoffmann-La Roche Ltd., Pharma Division, Basel, Switzerland.
Pöschl
B
B
Oesterhelt
G
G
Schönfeld
H J
HJ
Fountoulakis
M
M
D000596:Amino Acids / Q000032:analysis* / Q000737:chemistry; D001224:Aspartic Acid / Q000032:analysis / Q000737:chemistry*; D002849:Chromatography, Gas / Q000379:methods; D018698:Glutamic Acid / Q000032:analysis / Q000737:chemistry*; D005990:Glycerol / Q000737:chemistry* / Q000378:metabolism; D006851:Hydrochloric Acid / Q000737:chemistry; D006868:Hydrolysis; D013058:Mass Spectrometry / Q000379:methods; D008698:Mesylates / Q000737:chemistry; D011506:Proteins / Q000032:analysis / Q000737:chemistry*; D012808:Sigma Factor / Q000737:chemistry
D000596:Amino Acids; D008698:Mesylates; D011506:Proteins; D012808:Sigma Factor; C045880:methanesulfonic acid; D001224:Aspartic Acid; D018698:Glutamic Acid; D005990:Glycerol; D006851:Hydrochloric Acid