The Genetic Code tRNA’s charged with uncharged tRNA’s an amino acid Polypeptide chain Ribosome: note there are two A sites for binding a charged tRNA P (A-site and P-site) mRNA Steps in polypeptide elongation: 1. Prior to peptide bond formation: Peptidyl tRNA (bound to growing peptide chain) occupies the P-site. A charged tRNA occupies the A-site. 2. Peptide bond formation : Formation of bond is catalyzed by ribosome, and involves transfer of polypeptide from tRNA in the P-site to the tRNA in the A-site. This is stage shown above. 3. Translocation : Ribosome moves one codon down the mRNA. After this translocation the peptidyl tRNA will be in the P-site and the uncharged tRNA leaves the ribosomal complex. 1
anticodon codon A A A U U U mRNA 5 ’ 3 ’ Relationship between RNA and amino acid sequence: RNA polymer: 4 subunits / Polypeptide: 20 subunits • 1:1 can’t work • 2:1 provides 4 2 = 16 possibilities (still can’t work) • 3:1 provides 4 3 = 64 possibilities (BINGO!) 2
“Universal” genetic code Redundancy of genetic code Ala GCU, GCC, GCA, GCG Leu UUA, UUG, CUU, CUC, CUA, CUG Arg CGU, CGC, CGA, CGG, AGA, AGG Lys AAA, AAG Asn AAU, AAC Met AUG Asp GAU, GAC Phe UUU, UUC Cys UGU, UGC Pro CCU, CCC, CCA, CCG Gln CAA, CAG Ser UCU, UCC, UCA, UCG, AGU,AGC Glu GAA, GAG Thr ACU, ACC, ACA, ACG Gly GGU, GGC, GGA, GGG Trp UGG His CAU, CAC Tyr UAU, UAC Ile AUU, AUC, AUA Val GUU, GUC, GUA, GUG Start AUG, GUG Stop UAG, UGA, UAA 3
Features of the Genetic code: • The code is a triplet code. • The code is comma free • The code in non-overlapping • The code is highly degenerate • Wobble occurs in the anti-codon Pairing for leucine tRNA between standard Wobble pairing for Glycine tRNA is associated with the non- nucleotides standard nucleotide Inosine (I). Leu Leu Gly Gly Gly C C I C C I C C I G A G G A G G G U G G C G G A C U C C U U Codon – anticodon paring in the genetic code Wobble: non Watson-Crick bases pairing between codon and anti- codon. Wobble accounts for the ability of a single tRNA to recognise 5’ anticodon 3’ codon more than one codon. Wobble is predominantly a third codon G U or C positions effect, Some have suggested that the pattern of degeneracy C G of the genetic code is a adaptation to minimize errors arising from A U “unintended” wobble at third positions. U A or G I A, U or C 4
The genetic code determines how random changes to a protein-coding gene brought about by the process of mutation will impact the function of the encoded protein. 1. Degeneracy 2. Physiochemical similarity Evolution of the genetic code : How old is the genetic code ? • The earth is 4.6 billion years old • Youngest estimate of photosynthesis is 2.5 billion years ago Banded iron formations indicate the existence of free O 2 Photosynthesis evolved once in the cyanobacteria. It was later “stolen” by the eukaryotic ancestors of plants; in that it was probably acquired via the evolution of symbiosis between a eukaryotic cell and an cyanobacterium. Photosynthesis appears later in other lineages of eukaryotes as they “stole” it from each other. The cellular machinery for photosynthesis is highly derived and dependent upon a triplet genetic code. Hence the modern form of the code must have evolved prior to photosynthesis. Window: 4.6 billion - 2.5 billion years ago 5
Evolution of the genetic code : How old is the genetic code ? “Tree of life” showing three “domains, as estimated from rRNA sequences. Note that the Eucarya notion of a tree for the early divergences of life is Archaea controversial. I have also specified a basal trichotomy, as the inferred position of the root also is controversial. Archaea : prokaryotes with a plasma membrane of isoprene ester lipids. They have distinct acrhaebacterial-type riboosomes Bacteria: prokaryotes with a plasma membrane of fatty acid ester lipids. They have distinct Eubacterial--type ribosomes Eukarya: Cells with true nucleus. The plasma membrane consists of isoprene ester lipids. Ribosomes appear related to the DNA & RNA acrhaebacterial-type. Posess World double membrane bound organelles derived from bacterial endosymbionts RNA World DNA evolves Bacteria Genetic code evolves back here Eubacteria, and the genetic code are alder than 2.5 billion years, be we don’t know how much older. Evolution of the genetic code : How old is the genetic code ? Conditions for pre-cellular life (RNA World, etc) were not present prior to 3.9 biollion years ago. RNA world : • Pre-life, organic world • All RNA molecules • RNA acts as first genetic molecule and as enzyme • Controversial: RNA is unstable and known enzymatic activities are narrow Window: 3.9 billion - 2.5 billion years ago The genetic code is ANCIENT! 6
Evolution of the genetic code : How did it evolve ? 1. “Frozen accident” hypothesis: • Francic Crick (1968) • Code evlved very early • Most extreme version states that organization of the code was an accidient of history [i.e., neutral], and it have remianed unchanges ever since [frozen] • Less extreme verion lets natural selection play a role in optimizing the original code, then it becomes frozen Evolution of the genetic code : How did it evolve ? 2. Stereochemical compatibility hypothesis: One possible model for the evolution of tRNAs and the genetic code from a direct interaction between the involved amino acid and an “RNA handle” molecule. A. Early stage: direct interactions with “RNA handles” Amino acid with multiple codons in direct interaction RNA handle among many RNA Handles Peptide synthesis via catalytic RNA? Perhaps an additional catalytic RNA (Ribozyme) is required here? B. Later stage: use of a template and other RNA The use of a template molecules. RNA for some of the codons evolves. Modern tRNA structure evolves C. Late stage: RNA handles evolve into modern tRNAs RNA 7
Note: current tRNAs require an enzyme to catalyze cobalent bond between amino acid and tRNA Modern tRNAs have only very weak affinity for cognate amino acid Evolution of the genetic code : How did it evolve ? 3. Co-evolution with biosynthesis pathways 4. Minimize impact of mutation • degeneracy at third positons • physiochemical similarities Adapted from Knight et al. (1999) TIBS 24:241-247 5. Genetic flexibility 8
Evolution of the genetic code : Is the code frozen? Evolution of the genetic code : Is the code nearly frozen because of cellular constraints? Natural triplet codon in E. coli Artificial quadruplet codon in E. coli Andersen et al. PNAS (2004); and artificial Un-natural amino acid Natural amino acid quadruplet genetic code archaeal tRNALys archaeal tRNALys UCCU AGGA 9
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