Пирролизин: Құрамы, синтезі және метандық археялардағы рөлі

Pyrrolysine (symbol Pyl or O; encoded by the 'amber' stop codon UAG) is an α amino acid that is used in the biosynthesis of proteins in some methanogenic archaea and bacteria; it is not present in humans. It contains an α amino group (which is in the protonated –NH3+ form under biological conditions), a carboxylic acid group (which is in the deprotonated –COO− form under biological conditions). Its pyrroline side chain is similar to that of lysine in being basic and positively charged at neutral pH.

Nearly all genes are translated using only 20 standard amino acid building blocks. Two unusual genetically encoded amino acids are selenocysteine and pyrrolysine. Pyrrolysine was discovered in 2002 at the active site of methyltransferase enzyme from a methane producing archeon, Methanosarcina barkeri. and MALDI mass spectrometry, pyrrolysine is made up of 4 methylpyrroline 5 carboxylate in amide linkage with the εN of lysine.

Pyrrolysine is synthesized in vivo by joining two molecules of L lysine. One molecule of lysine is first converted to (3R) 3 methyl D ornithine, which is then ligated to a second lysine. An NH2 group is eliminated, followed by cyclization and dehydration step to yield L pyrrolysine.

The extra pyrroline ring is incorporated into the active site of several methyltransferases, where it is believed to rotate relatively freely. It is believed that the ring is involved in positioning and displaying the methyl group of methylamine for attack by a corrinoid cofactor. The proposed model is that a nearby carboxylic acid bearing residue, glutamate, becomes protonated, and the proton can then be transferred to the imine ring nitrogen, exposing the adjacent ring carbon to nucleophilic addition by methylamine. The positively charged nitrogen created by this interaction may then interact with the deprotonated glutamate, causing a shift in ring orientation and exposing the methyl group derived from the methylamine to the binding cleft where it can interact with corrinoid. In this way a net CH3+ is transferred to the cofactor's cobalt atom with a change of oxidation state from I to III. The methylamine derived ammonia is then released, restoring the original imine. For example, the system provided one of two fluorophores incorporated site specifically within calmodulin to allow the real time examination of changes within the protein by FRET spectroscopy, and site specific introduction of a photocaged lysine derivative. (See Expanded genetic code)

It was originally proposed that a specific downstream sequence "PYLIS", forming a stem loop in the mRNA, forced the incorporation of pyrrolysine instead of terminating translation in methanogenic archaea. This would be analogous to the SECIS element for selenocysteine incorporation.

The pylT (tRNA) and pylS (aa tRNA synthase) genes are part of an operon of Methanosarcina barkeri, with homologues in other sequenced members of the Methanosarcinaceae family: M. acetivorans, M. mazei, and M. thermophila. Pyrrolysine containing proteins are known to include monomethylamine methyltransferase (mtmB), dimethylamine methyltransferase (mtbB), and trimethylamine methyltransferase (mttB). Homologs of pylS and pylT have also been found in an Antarctic archaeon, Methanosarcina barkeri and a Gram positive bacterium, Desulfitobacterium hafniense. The other genes of the Pyl operon mediate pyrrolysine biosynthesis, leading to description of the operon as a "natural genetic code expansion cassette". A number of evolutionary scenarios have been proposed for the pyrrolysine system. The current (2022) view, given available sequences for tRNA and Pyl tRNA (PylRS) synthase genes, is that:
tRNA(Pyl) diverged from tRNA(Phe) some time between the divergence of the three domains (~LUCA) and the divergence of archaeal phyla, but was lost in non archaeal lineages; It is now known that a wide range of prokaryotes have these two genes. It is now known that some non methanogens also have these two genes, but the dating was not too far off. This is probably true based on the 2022 study, though the paper originally assumed a link to methanogenesis. Further study found that the genes encoding LysRS1 and LysRS2 are not required for normal growth on methanol and methylamines with normal methyltransferase levels, and they cannot replace pylS in a recombinant system for UAG amber stop codon suppression.