Directed Evolution of a Model Primordial Enzyme Provides Insights into the Development of the Genetic Code

English version České info

The contemporary proteinogenic repertoire contains 20 amino acids with diverse functional groups and side chain geometries. Primordial proteins, in contrast, were presumably constructed from a subset of these building blocks. Subsequent expansion of the proteinogenic alphabet would have enhanced their capabilities, fostering the metabolic prowess and organismal fitness of early living systems. While the addition of amino acids bearing innovative functional groups directly enhances the chemical repertoire of proteomes, the inclusion of chemically redundant monomers is difficult to rationalize. Here, we studied how a simplified chorismate mutase evolves upon expanding its amino acid alphabet from nine to potentially 20 letters. Continuous evolution provided an enhanced enzyme variant that has only two point mutations, both of which extend the alphabet and jointly improve protein stability by >4 kcal/mol and catalytic activity tenfold. The same, seemingly innocuous substitutions (Ile→Thr, Leu→Val) occurred in several independent evolutionary trajectories. The increase in fitness they confer indicates that building blocks with very similar side chain structures are highly beneficial for fine-tuning protein structure and function.

Vyšlo v časopise: Directed Evolution of a Model Primordial Enzyme Provides Insights into the Development of the Genetic Code. PLoS Genet 9(1): e32767. doi:10.1371/journal.pgen.1003187
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.pgen.1003187

Souhrn

Zdroje

1. JoyceGF (1989) RNA evolution and the origins of life. Nature 338: 217–224.

2. CrickFHC (1968) The origin of the genetic code. J Mol Biol 38: 367–379.

3. OrgelLE (1968) Evolution of the genetic apparatus. J Mol Biol 38: 381–393.

4. NarlikarGJ, HerschlagD (2003) Mechanistic aspects of enzymatic catalysis: Lessons from comparison of RNA and protein enzymes. Annu Rev Biochem 66: 19–59.

5. DoudnaJA, LorschJR (2005) Ribozyme catalysis: Not different, just worse. Nat Struct Mol Biol 12: 395–402.

6. WongJT-F (2005) Coevolution theory of the genetic code at age thirty. BioEssays 27: 416–425.

7. MillerSL (1953) A production of amino acids under possible primitive earth conditions. Science 117: 528–529.

8. KvenvoldenK, LawlessJ, PeringK, PetersonE, FloresJ, et al. (1970) Evidence for extraterrestrial amino-acids and hydrocarbons in the Murchison meteorite. Nature 228: 923–926.

9. SicheriF, YangDSC (1995) Ice-binding structure and mechanism of an antifreeze protein from winter flounder. Nature 375: 427–431.

10. RiddleDS, SantiagoJV, Bray-HallST, DoshiN, GrantcharovaVP, et al. (1997) Functional rapidly folding proteins from simplified amino acid sequences. Nat Struct Mol Biol 4: 805–809.

11. TanakaJ, YanagawaH, DoiN (2011) Comparison of the frequency of functional SH3 domains with different limited sets of amino acids using mRNA display. PLoS ONE 6: e18034 doi:10.1371/journal.pone.0018034.

12. AkanumaS, KigawaT, YokoyamaS (2002) Combinatorial mutagenesis to restrict amino acid usage in an enzyme to a reduced set. Proc Natl Acad Sci USA 99: 13549–13553.

13. MacBeathG, KastP, HilvertD (1998) A small, thermostable, and monofunctional chorismate mutase from the archaeon Methanococcus jannaschii. Biochemistry 37: 10062–10073.

14. WalterKU, VamvacaK, HilvertD (2005) An active enzyme constructed from a 9-amino acid alphabet. J Biol Chem 280: 37742–37746.

15. TrifonovEN (2000) Consensus temporal order of amino acids and evolution of the triplet code. Gene 261: 139–151.

16. KamtekarS, SchifferJ, XiongH, BabikJ, HechtM (1993) Protein design by binary patterning of polar and nonpolar amino acids. Science 262: 1680–1685.

17. FisherMA, McKinleyKL, BradleyLH, ViolaSR, HechtMH (2012) De novo designed proteins from a library of artificial sequences function in Escherichia coli and enable cell growth. PLoS ONE 6: e15364 doi:10.1371/journal.pone.0015364.

18. TaylorSV, WalterKU, KastP, HilvertD (2001) Searching sequence space for protein catalysts. Proc Natl Acad Sci USA 98: 10596–10601.

19. KastP, Asif-UllahM, JiangN, HilvertD (1996) Exploring the active site of chorismate mutase by combinatorial mutagenesis and selection: The importance of electrostatic catalysis. Proc Natl Acad Sci USA 93: 5043–5048.

20. CambrayG, MazelD (2008) Synonymous genes explore different evolutionary landscapes. PLoS Genet 4: e1000256 doi:10.1371/journal.pgen.1000256.

21. NeuenschwanderM, ButzM, HeintzC, KastP, HilvertD (2007) A simple selection strategy for evolving highly efficient enzymes. Nat Biotechnol 25: 1145–1147.

22. MutzelR, MarlièreP (2000) Method and device for selecting accelerated proliferation of living cells in suspension. WO2000034433.

23. MacBeathG, KastP, HilvertD (1998) Exploring sequence constraints on an interhelical turn using in vivo selection for catalytic activity. Protein Sci 7: 325–335.

24. StewartJ, WilsonDB, GanemB (1990) A genetically engineered monofunctional chorismate mutase. J Am Chem Soc 112: 4582–4584.

25. MacBeathG, KastP (1998) UGA read-through artifacts - when popular gene expression systems need a pATCH. BioTechniques 24: 789–794.

26. ZhangY (2008) I-TASSER server for protein 3D structure prediction. BMC Bioinf 9: 40.

27. LeeAY, KarplusPA, GanemB, ClardyJ (1995) Atomic structure of the buried catalytic pocket of Escherichia coli chorismate mutase. J Am Chem Soc 117: 3627–3628.

28. DillKA, BrombergS, YueK, ChanHS, FtebigKM, et al. (1995) Principles of protein folding - A perspective from simple exact models. Protein Sci 4: 561–602.

29. WolynesPG (1997) As simple as can be? Nat Struct Biol 4: 871–874.

30. RogersJ, JoyceGF (1999) A ribozyme that lacks cytidine. Nature 402: 323–325.

31. ReaderJS, JoyceGF (2002) A ribozyme composed of only two different nucleotides. Nature 420: 841–844.

32. RogersJ, JoyceGF (2001) The effect of cytidine on the structure and function of an RNA ligase ribozyme. RNA 7: 395–404.

33. YooTH, LinkAJ, TirrellDA (2007) Evolution of a fluorinated green fluorescent protein. Proc Natl Acad Sci USA 104: 13887–13890.

34. LiuCC, MackAV, TsaoM-L, MillsJH, LeeHS, et al. (2008) Protein evolution with an expanded genetic code. Proc Natl Acad Sci USA 105: 17688–17693.

35. BacherJM, HughesRA, WongJT-F, EllingtonAD (2004) Evolving new genetic codes. Trends Ecol Evol 19: 69–75.

36. LemeignanB, SonigoP, MarlièreP (1993) Phenotypic suppression by incorporation of an alien amino acid. J Mol Biol 231: 161–166.

37. GamperM, KastP (2005) Strategy for chromosomal gene targeting in RecA-deficient Escherichia coli strains. BioTechniques 38: 405–408.

38. OostenbrinkC, VillaA, MarkAE, van GunsterenWF (2004) A biomolecular force field based on the free enthalpy of hydration and solvation: The GROMOS force-field parameter sets 53A5 and 53A6. J Comput Chem 25: 1656–1676.

39. OostenbrinkC, SoaresTA, van der VegtNFA, van GunsterenWF (2005) Validation of the 53A6 GROMOS force field. Eur Biophys J 34: 273–284.

40. ChristenM, HünenbergerPH, BakowiesD, BaronR, BürgiR, et al. (2005) The GROMOS software for biomolecular simulation: GROMOS05. J Comput Chem 26: 1719–1751.

41. EichenbergerAP, AllisonJR, DolencJ, GeerkeDP, HortaBAC, et al. (2011) GROMOS++ software for the analysis of biomolecular simulation trajectories. J Chem Theory Comput 7: 3379–3390.

42. LiuDR, CloadST, PastorRM, SchultzPG (1996) Analysis of active site residues in Escherichia coli chorismate mutase by site-directed mutagenesis. J Am Chem Soc 118: 1789–1790.

43. SassoS, RamakrishnanC, GamperM, HilvertD, KastP (2005) Characterization of the secreted chorismate mutase from the pathogen Mycobacterium tuberculosis. FEBS J 272: 375–389.

44. MatteiP, KastP, HilvertD (1999) Bacillus subtilis chorismate mutase is partially diffusion-controlled. Eur J Biochem 261: 25–32.

45. GamperM, HilvertD, KastP (2000) Probing the role of the C-terminus of Bacillus subtilis chorismate mutase by a novel random protein-termination strategy. Biochemistry 39: 14087–14094.