Diphthamide was named after diphtheria, an infectious disease caused by Corynebacterium diptheriae. Diphtheria toxin attaches an ADP-ribose fragment to elongation factor eEF2 and this inhibits protein synthesis and kills the target cells. eEF2 normally splits GTP and uses the energy released to move the peptidyl-tRNA from the A-site to the P-site. ADP-ribosylated eEF2 still binds GTP but cannot hydrolyze it or translocate the peptidyl-tRNA.

Very rarely, the stop codon UGA is read as the unusual amino acid selenocysteine.

The stop codon UAG is occasionally translated as the rare amino acid, pyrrolysine.

and proteins involved has shown that selenocysteine is encoded by UGA. However, UGA is one of the stop codons. Apparently, UGA is normally read as "stop" but is occasionally translated to give selenocysteine, which therefore has the honor of being the 21st genetically encoded amino acid.The choice between "stop" and selenocysteine depends on a special recognition sequence in the following part of the gene—the selenocysteine insertion sequence (SECIS element). Selenocysteine has its own tRNA and a special protein factor to escort charged tRNA-Sec to the ribosome. In fact, selenocysteine-tRNA is initially charged with serine. Then the attached serine is enzy-matically modified to form selenocysteine.

When bacteria use selenocysteine, the selenocysteine insertion sequence forms a stem and loop structure in the mRNA molecule just after the UGA. SelB protein recognizes both charged tRNA-Sec and this stem and loop. Thus selenocysteine bound to tRNA is delivered to the right place (Fig. 8.30A). [In bacteria, the stem and loop form temporarily from part of the coding sequence, and this section of the mRNA is therefore translated after insertion of the selenocysteine.] In mammals, the stem and loop structure is found beyond the end of the coding sequence, in the 3'-untranslated region—not next to the critical UGA codon! A pair of proteins is responsible for binding the tRNA-Sec and recognizing stem and loop. Somehow, they deliver the tRNA-Sec to the correct position for insertion (Fig. 8.30B).

Selenocysteine is an analog of cysteine,but has selenium instead of sulfur (Fig. 8.31). Selenium is more susceptible to oxidation than sulfur and so proteins that contain it must be protected from oxygen. Examples are the formate dehydrogenases found in many bacteria. These contain selenocysteine in their active sites and function in anaerobic metabolism. They are inactivated by oxygen and are normally made only in the absence of air. Higher organisms contain about 20 proteins that contain selenocysteine. Zebrafish selenoprotein P contains 17 Sec residues, the largest number in any known protein.

Pyrrolysine: The 22nd Amino Acid

In 2002, a 22nd genetically encoded amino acid was discovered—pyrrolysine, a derivative of lysine with an attached pyrroline ring (Fig. 8.32). This is found in a few archaebacteria where it is encoded by the stop codon UAG in occasional proteins. Pyrrolysine was first discovered in the active site of methylamine methyl-transferases found in methane producing archaebacteria of the genus Methanosarcina. An unusual selenocyteine insertion sequence (SECIS element) Recognition sequence that signals for insertion of selenocysteine at a UGA stop codon

Pyrrolysine: The 22nd Amino Acid 229

FIGURE 8.30 Stop Codon

A) In bacteria, the tRNA carrying selenocysteine (Sec) first binds to SelB and the complex then binds to a stem and loop in the mRNA. This aligns the tRNASec with a UGA codon within the coding sequence on the mRNA. Selenocysteine is then inserted as part of the growing polypeptide. Only the fully bound complex is shown. B) In mammals, the protein that binds the stem and loop and the tRNASec is called eEFsec. In addition, the stem and loop are more distant, being found after the stop codon.

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