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Simulation Results

Simulation Results. Mutation results for MG1655 and O157:H7 E-Coli strains. Detected Translation Signals. MG1655. Detected translation signals. -1. -2. -3. Termination. Codon. Initiation Codon. -4. Average Free Energy (kcal/mol). -5. -6. -7. Shine Dalgarno. Signal. -8.

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Simulation Results

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  1. Simulation Results Mutation results for MG1655 and O157:H7 E-Coli strains

  2. Detected Translation Signals MG1655 Detected translation signals -1 -2 -3 Termination Codon Initiation Codon -4 Average Free Energy (kcal/mol) -5 -6 -7 Shine Dalgarno Signal -8 Coding (translated) Non coding (arbitrary) -9 0 50 100 150 200 250 300 350 400 450 500 Position

  3. Detected Translation Signals O157:H7

  4. Mutations • Experimental results obtained by mutating regions of the 3’ end of the 16SrRNA are compared with results obtained by incorporating these mutations in the 16SrRNA based codebook of our model. • Jacob introduced a point mutation in the 5th position of the 16SrRNA [16]. Specifically, the 5th position in the arrangement illustrated below: Original UAAGGAGGUGAUC Mutated UAACCUCCUGAUC • This point mutation consisted in a change of the nucleotide C → U in the ribosome small subunit. This is equivalent to make a mutation from G → A in the complement sequence shown above. The result of this mutation was a reduction in the level of protein synthesis. Fig. 1 and Fig. 2 show how the recognition of the Shine-Dalgarno signal is affected for the Jacob mutation. It can be inferred from the plot that the levels of protein production will be reduced but not completely stopped.

  5. MG1655 Fig 1

  6. O157:H7 Partial Loss due to mutation Fig 2

  7. Hui and De Boer did another published record of the behavior of the protein synthesis under mutations in the 3’ end of the 16SrRNA. • In this experiment, the mutations were done in positions 4 to 8(GGAGG → CCUCC) and positions 5 to 7 (GAG → UGU). Original UAAGGAGGUGAUC Mutated 1 UAACCUCCUGAUC Mutated 2 UAACUGUCUGAUC • The results of both mutations were lethal for the organism in the sense that the production of proteins stopped. After introducing these two mutations, the results showed a complete loss of the SD signal. Hence, it can be inferred that the translation will never take place. This is illustrated in Fig. 3 and Fig. 4.

  8. MG1655 Fig 3

  9. O157:H7 Fig 4

  10. Point Mutations • To exploit the model further, point mutations have been introduced in all positions of the last 13 bases of the 16SrRNA molecule in order to study their influence on the process of translation. • The obtained results are summarized in Table I by quantizing into 5 levels the influence of these mutations on each of the translation signals (SD, initiation, stop). The levels are: – represents no influence in the recognition of the signal,  represents a strong negative influence ↓ a weak influence ↑ a weak positive influence  a strong positive influence.

  11. Table I: POINT MUTATIONS IN 16SrRNA For example, results show how a mutation in position 5 has a strong negative influence in the recognition of the SD signal, just as found in the Jacob investigation.

  12. Remarkable Findings • A mutation in position 8 has no influence in the detection of the translation signals, probably the reason is that the role of this nucleotide is to introduce spacing at the moment of decoding the mRNA sequence. • A mutation at position 6 has nearly the same influence as a mutation at position 5. • A mutation at position 9 affects the recognition of the initiation codon even if it does not affect the SD signal. This could lead to a wrong initiation of translation or a “frame shift”. • Exactly the central part of the 13 bases (bases 4-8) which influences the SD is missing in eukaryotes. The rest of the sequence that involves AUG and stop codon recognition are still there.

  13. MG1655 1st Position Mutation in the 1st Position (U to A) (SD ─ , IC ─ , SC ↓ )

  14. O157:H7 1st Position Mutation in the 1st Position (U to A) (SD ─ , IC ─ , SC ↓ )

  15. MG1655 1st Position Mutation in the 1st Position (U to A) (SD ─ , IC ─ , SC ↓ )

  16. O157:H7 1st Position Mutation in the 1st Position (U to A) (SD ─ , IC ─ , SC ↓ )

  17. MG1655 2nd Position Mutation in the 2nd Position (U to A) (SD ─ , IC ─ , SC  )

  18. O157:H7 2nd Position Mutation in the 2nd Position (U to A) (SD ─ , IC ─ , SC  )

  19. MG1655 2nd Position Mutation in the 2nd Position (U to A) (SD ─ , IC ─ , SC  )

  20. O157:H7 2nd Position Mutation in the 2nd Position (U to A) (SD ─ , IC ─ , SC  )

  21. MG1655 3rd Position Mutation in the 3rd position (A to U) (SD ─ , IC , SC ↓ )

  22. O157:H7 3rd Position Mutation in the 3rd position (A to U) (SD ─ , IC , SC ↓ )

  23. MG1655 3rd Position Mutation in the 3rd position (A to U) (SD ─ , IC , SC ↓ )

  24. O157:H7 3rd Position Mutation in the 3rd position (A to U) (SD ─ , IC , SC ↓ )

  25. MG1655 4th Position Mutation in the 4th position (G to C) (SD ↓ , IC , SC ─ )

  26. O157:H7 4th Position Mutation in the 4th position (G to C) (SD ↓ , IC , SC ─ )

  27. MG1655 4th Position Mutation in the 4th position (G to C) (SD ↓ , IC , SC ─ )

  28. O157:H7 4th Position Mutation in the 4th position (G to C) (SD ↓ , IC , SC ─ )

  29. MG1655 5th Position Mutation in the 5th position (G to C) (SD  , IC↓ , SC ─ )

  30. O157:H7 5th Position Mutation in the 5th position (G to C) (SD  , IC↓ , SC ─ )

  31. MG1655 5th Position Mutation in the 5th position (G to C) (SD  , IC↓ , SC ─ )

  32. O157:H7 5th Position Mutation in the 5th position (G to C) (SD  , IC↓ , SC ─ )

  33. MG1655 6th Position Mutation in the 6th position (A to U) (SD  , IC↓ , SC ─ )

  34. O157:H7 6th Position Mutation in the 6th position (A to U) (SD  , IC↓ , SC ─ )

  35. MG1655 6th Position Mutation in the 6th position (A to U) (SD  , IC↓ , SC ─ )

  36. O157:H7 6th Position Mutation in the 6th position (A to U) (SD  , IC↓ , SC ─ )

  37. MG1655 7th Position Mutation in the 7th position (G to C) (SD ↓ , IC↓ , SC ─ )

  38. O157:H7 7th Position Mutation in the 7th position (G to C) (SD ↓ , IC↓ , SC ─ )

  39. MG1655 7th Position Mutation in the 7th position (G to C) (SD ↓ , IC↓ , SC ─ )

  40. O157:H7 7th Position Mutation in the 7th position (G to C) (SD ↓ , IC↓ , SC ─ )

  41. MG1655 8th Position Mutation in the 8th position (G to C) (SD ─ , IC ─ , SC ─ )

  42. O157:H7 8th Position Mutation in the 8th position (G to C) (SD ─ , IC ─ , SC ─ )

  43. MG1655 8th Position Mutation in the 8th position (G to C) (SD ─ , IC ─ , SC ─ )

  44. O157:H7 8th Position Mutation in the 8th position (G to C) (SD ─ , IC ─ , SC ─ )

  45. MG1655 9th Position Mutation in the 9th position (U to A) (SD ─ , IC , SC ↓ )

  46. O157:H7 9th Position Mutation in the 9th position (U to A) (SD ─ , IC , SC ↓ )

  47. MG1655 9th Position Mutation in the 9th position (U to A) (SD ─ , IC , SC ↓ )

  48. O157:H7 9th Position Mutation in the 9th position (U to A) (SD ─ , IC , SC ↓ )

  49. MG1655 10th Position Mutation in the 10th position (U to A) (SD ─ , IC ↓ , SC  )

  50. O157:H7 10th Position Mutation in the 10th position (U to A) (SD ─ , IC ↓ , SC  )

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