IGEM:Stanford/2009/Notebook/Marys iGEM Notebook/2009/08/20

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Fitting of 5-methyl-trp and L-trp Binding with Wild Type Repressor <Val58>

The reason why we do this fitting is because the set of fitting results for L-trp + wild type repressor doesn't fit in as well for other situations, including trp binding mutant repressor, 5MT binding with wild type or mutant repressor. As shown here:4 different situations of binding

The following analysis is based the on assumption that k_R value for 5MT and trp, in each one of the three different models(1-step,2-step,3-step binding of repressor and operator), are the same.

Since we're lacking of dissociation constant of a single 5MT binding with one single site of the repressor (R) dimer, the binding process of trp/5MT + R is considered as a one-step process in the following plotting and analysis. The value of k_T (dissociation constant of 2 molecules of trp or 5MT binding to aporepressor dimer) are from papers.

The reason why I added in another "2-step binding" model is because, as discussed in some papers, and binding affinity of one of the three binding sites of the operator is really weak. And Reference [1] actually used the 2-step binding model. So I tried it out for comparison.

Why none of the three models fit both trp and 5MT perfectly? It's probably due to the following reasons:
  1. Inaccurate models
  2. Inaccurate k_T values (Plus we're actually implying a one-step trp+repressor model here. It's inaccurate in itself.)
  3. Inaccurate k_c values (The k_coupling value is set as 15 in both 2-step and 3-step binding models. kc=40+_15, as shown in [1]. But we're not quite sure it fits with our model.)


References

  1. In vivo and in vitro Studies of TrpR-DNA Interactions. Jie Yang, Angelo Gunasekera†, Teresa A. Lavoie, Lihua Jin, Dale E. A. Lewis and Jannette Carey. J. Mol. Biol. (1996) 258, 37–52

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