I'm trying to obtain a better understanding of the Guthrie Test, which checks whether people have a disease called PKU. The Guthrie test uses bacteria to check for Phenylalanine (the amino acid) in blood samples. If both Phe and B-2-thienylalanine are available on the medium, the bacteria will grow. However, if only B-2-thienylalanine is available the bacteria will not grow.

In all of the articles I can find, I have seen no explanation, just a link to a paper that I can't find on the internet.: " A standard culture of Bacillus subtilis was incubated on agar in the presence of an antagonist of phenylalanine (B-2-thienylalanine), which prevents the bacteria from growing.3 " Link The reference they mention here is: "The inhibition assay: Its use in screening urinary specimens for metabolic differences associated with mental retardation" by R. Guthrie from 1960.

Do we know what mechanism this protein is inhibiting?

Any help would be greatly appreciated.


1 Answer 1


β-2-thienylalanine is an analogue of phenylalanine. It is incorporated into proteins in place of phenylalanine, and the resulting proteins are inactive because of the substitution. Consequently growth medium supplemented with β-2-thienylalanine will not support bacterial growth.

Addition of phenylalanine to a medium containing β-2-thienylalanine will result in a competition between the two compounds - if there is an excess of phenylalanine then this will out-compete the analogue and incorporation of the analogue will be minimised, allowing growth. Blood from a patient with PKU contains phenylalanine, so the addition of this to the growth medium will relieve inhibition of growth.

Here is a link to a patent application in which a screen for E. coli strains overproducing phenylalanine was based upon the same principle.

  • $\begingroup$ Thanks Dr. Boyd! I greatly, greatly appreciate your help! $\endgroup$
    – Sean
    Nov 5, 2013 at 19:44

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