4 things to remember about Amino Acid Analysis of proteins & peptides

Amino Acid analysis
Sep 23. 2016

Why perform Amino Acid Analysis?

Amino acid analysis of proteins is a method to determine the absolute amounts of individual amino acids in a sample. The method can be applied to samples containing free amino acids. However it may also be used on peptides and protein samples after hydrolysis into amino acids [1, 2].

AA analysis can be used for determination of the relative composition of amino acids in a protein. It can also be used for determination of the absolute amount of a protein or peptide if the amino acid sequence is known. Finally, it is used for purity estimation of a purified protein [3].

In amino acid analysis it should be noted that:

  1. Serine and threonine are degraded slightly during acid hydrolysis, and recoveries can be 10% lower than expected.
  2. Methionine can be oxidized during hydrolysis, usually less than 10% is oxidized.
  3. Valine and isoleucine bonds (Val-Val, Ile-Val, Val-Ile, Ile-Ile) are difficult to hydrolyse and recoveries can be 5-15% lower than expected.
  4. Glycine content is often higher than expected because it is a frequent contaminant due to its use in many buffers. Analysis of known amounts of AA standards is used to determine a compensation factor to correct for differences in ninhydrin reactivity [1-3].

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[1]          Rutherfurd et al: “Amino acid analysis“, Current Protocols in Protein Science2009

[2]          Rutherfurd et al: “Quantitative amino acid analysis“, Current Protocols in Protein Science2011

[3]          Noble et al: “Quantification of protein“, Methods in Enzymology2009

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