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Pyrrolysine Information

Pyrrolysine is a naturally occurring, genetically coded amino acid used by some methanogenic archaea and one known bacterium in enzymes that are part of their methane-producing metabolism. It is similar to lysine, but with an added pyrroline ring linked to the end of the lysine side chain. Produced by a specific tRNA and aminoacyl tRNA synthetase, it forms part of an unusual genetic code in these organisms, and is considered the 22nd proteinogenic amino acid.

The joint nomenclature committee of the IUPAC/IUBMB has officially recommended the three-letter symbol Pyl and the one-letter symbol O for pyrrolysine.

Contents

Introduction and context

One key function of the genome is to direct production of proteins using genetic sequences that determine when or if each protein will be produced; what cells will produce it; and where it is located in the cell. Proteins form much of the physical structure of the body and catalyze a wide variety of chemical reactions, giving the genome the ability to control the body's biochemistry. Nearly all proteins are made using only 20 standard building blocks called amino acids, which are often assembled in very long sequences according to a standard genetic code. Specialized chemical reactions often require alterations of proteins after the fact by posttranslational modification, or protein binding to specific cofactors. Yet the genetic code itself is exactly the same among very many organisms, so that when researchers sequence DNA from new or unknown sources they can often immediately draw conclusions about the chemical activity it carries out based on the assumption that a standard genetic code applies. The discovery of unusual amino acids specified by an expansion of the genetic code can call this assumption into question, so it is important to understand any such aberrations. Additionally, these variations indicate that the process of evolution that led to the establishment of the genetic code did not end before the universal common ancestor perhaps some three to four billion years ago, but remains accessible to study even in the present day.

Composition

As determined by X-ray crystallography[1] and MALDI mass spectrometry, pyrrolysine is made up of 4-methylpyrroline-5-carboxylate in amide linkage with the ϵN of lysine.[2]

Catalytic function

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 models 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.[1]

Genetic coding

Unlike posttranslational modifications of lysine such as hydroxylysine, methyllysine, and hypusine, pyrrolysine is incorporated during translation (protein synthesis) as directed by the genetic code, just like the 20 standard amino acids. It is encoded in mRNA by the UAG codon, which in most organisms is the 'amber' stop codon. This requires only the presence of the pylT gene, which encodes an unusual transfer RNA (tRNA) with a CUA anticodon, and the pylS gene, which encodes a class II aminoacyl-tRNA synthetase that charges the pylT-derived tRNA with pyrrolysine.

This novel tRNA-aaRS pair ("orthogonal pair") is independent of other synthetases and tRNAs in Escherichia coli, and further possesses some flexibility in the range of amino acids processed, making it an attractive tool to allow the placement of a possibly wide range of functional chemical groups at arbitrarily specified locations in modified proteins.[3][4] 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,[5] and site-specific introduction of a photocaged lysine derivative.[6] (See Expanded genetic code)

Evolution

The pylT and pylS 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 genes 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, Methanococcoides burtoni and a Gram-positive bacterium, Desulfitobacterium hafniense.[7][8]

The occurrence in Desulfitobacterium is of special interest, because bacteria and archaea are separate domains in the three-domain system by which living things are classified. When use of the amino acid appeared confined to the Methanosarcinaceae, the system was described as a "late archaeal invention" by which a 21st amino acid was added to the genetic code.[9] Afterward it was concluded that "PylRS was already present in the last universal common ancestor" some 3 billion years ago, but it only persisted in organisms using methylamines as energy sources.[10] Another possibility is that evolution of the system involved a horizontal gene transfer between unrelated microorganisms.[11] The other genes of the Pyl operon mediate pyrrolysine biosynthesis, leading to description of the operon as a "natural genetic code expansion cassette".[12]

Some differences exist between the bacterial and archaeal systems studied. Homology to pylS is broken into two separate proteins in D. hafniense. Most notably, the UAG codon appears to act as a stop codon in many of that organism's proteins, with only a single established use in coding pyrrolysine in that organism. By contrast, in methanogenic archaea it was not possible to identify any unambiguous UAG stop signal.[7] Because there was only one known site where pyrrolysine is added in D. hafniense it was not possible to determine whether some additional sequence feature, analogous to the SECIS element for selenocysteine incorporation, might control when pyrrolysine is added. It was previously 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. However, the PYLIS model has lost favor in view of the lack of structural homology between PYLIS elements and the lack of UAG stops in those species.

Potential for an alternate translation

The tRNA(CUA) can be charged with lysine in vitro by the concerted action of the M. barkeri Class I and Class II Lysyl-tRNA synthetases, which do not recognize pyrrolysine. Charging a tRNA(CUA) with lysine was originally hypothesized to be the first step in translating UAG amber codons as pyrrolysine, a mechanism analogous to that used for selenocysteine. More recent data favor direct charging of pyrrolysine on to the tRNA(CUA) by the protein product of the pylS gene, leading to the suggestion that the LysRS1:LysRS2 complex may participate in a parallel pathway designed to ensure that proteins containing the UAG codon can be fully translated using lysine as a substitute amino acid in the event of pyrrolysine deficiency.[13] 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.[14]

See also

References

External links

References

  1. ^ a b Bing Hao, Weimin Gong, Tsuneo K. Ferguson, Carey M. James, Joseph A. Krzycki, Michael K. Chan (2002-05-24). "A New UAG-Encoded Residue in the Structure of a Methanogen Methyltransferase". Science. pp. 1462–1466. doi:10.1126/science.1069556. http://www.sciencemag.org/cgi/content/full/296/5572/1462/F4.
  2. ^ Jitesh A. Soares, Liwen Zhang, Rhonda L. Pitsch, Nanette M. Kleinholz, R. Benjamin Jones, Jeremy J. Wolff, Jon Amster, Kari B. Green-Church, and Joseph A. Krzycki (2005-11-04). "The residue mass of L-pyrrolysine in three distinct methylamine methyltransferases". The Journal of biological chemistry (Journal of Biological Chemistry) 280 (44): 36962–36969. doi:10.1074/jbc.M506402200. PMID 16096277. http://www.jbc.org/content/280/44/36962.long.
  3. ^ PMID 15380192
  4. ^ PMID 19063902
  5. ^ PMID 19156778
  6. ^ PMID 19378306
  7. ^ a b Reviewed in Yan Zhang, Pavel V. Baranov, John F. Atkins, and Vadim N. Gladyshev (May 27, 2005). "Pyrrolysine and selenocysteine use dissimilar decoding strategies". Journal of Biological Chemistry 280 (21): 20740–20751. doi:10.1074/jbc.M501458200. PMID 15788401. http://www.jbc.org/content/280/21/20740.long.
  8. ^ Yan Zhang and Vadim N. Gladyshev (2007). "High content of proteins containing 21st and 22nd amino acids, selenocysteine and pyrrolysine, in a symbiotic deltaproteobacterium of gutless worm Olavius algarvensis". Nucleic Acids Research 35 (15): 4952–4963. doi:10.1093/nar/gkm514. PMID 17626042. PMC 1976440. http://nar.oxfordjournals.org/cgi/content/full/35/15/4952.
  9. ^ Alexandre Ambrogelly, Sarath Gundllapalli, Stephanie Herring, Carla Polycarpo, Carina Frauer and Dieter Söll (2007-02-27). "Pyrrolysine is not hardwired for cotranslational insertion at UAG codons". PNAS 104 (9): 3141–3146. doi:10.1073/pnas.0611634104. PMID 17360621. PMC 1805618. http://www.pnas.org/content/104/9/3141.long.
  10. ^ Kayo Nozawa, Patrick O’Donoghue, Sarath Gundllapalli, Yuhei Araiso, Ryuichiro Ishitani, Takuya Umehara, Dieter Soll, and Osamu Nureki (2009-02-26). "Pyrrolysyl-tRNA synthetase:tRNAPyl structure reveals the molecular basis of orthogonality". pp. 1163–1167. doi:10.1038/nature07611.. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2648862/pdf/nihms77751.pdf.
  11. ^ PMID 19271184
  12. ^ David G. Longstaff, Ross C. Larue, Joseph E. Faust, Anirban Mahapatra, Liwen Zhang, Kari B. Green-Church, and Joseph A. Krzycki (2007-01-16). "A natural genetic code expansion cassette enables transmissible biosynthesis and genetic encoding of pyrrolysine". Proc Natl Acad Sci U S A 104 (3): 1021–6. doi:10.1073/pnas.0610294104. PMID 17204561. PMC 1783357. http://www.pnas.org/content/104/3/1021.long.
  13. ^ "An aminoacyl-tRNA synthetase that specifically activates pyrrolysine". 2004-08-24. pp. 12450–12454. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC515082/.
  14. ^ PMID 17542922

Categories: Amino acids | Pyrrolines

 

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