User:Matt Hartings/Notebook/Photosynthesis/2013/10/03

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(Procedure and Data)
Current revision (14:17, 3 October 2013) (view source)
(Procedure and Data)
 

Current revision

Protein Re-engineering Main project page
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Objective

Take kinetic data for Mb catalyzed o-phenylenediamine + H2O2 reaction at higher temperatures (25, 35, and 45C)

Procedure and Data

Stock solutions

sample 1

  1. Stock solutions:
    1. Myoglobin
      1. 2.0mg (Mb:KCl lyophilized from Citrate, 5mg:500mg) in 1.0mL water
    2. OPD
      1. 4.1mg OPD in 2.4mL water
    3. H2O2
      1. 0.114mL 30% H2O2 in 0.887mL water
  2. Experimental Run
    1. Sample chamber set to 25C and to stir
    2. N2 run through the cuvette holder to reduce condensation on cuvette
    3. 2.025mL OPD stock solution
    4. 0.75mL H2O2 stock solution
      1. Start Data Collection
      2. 1ms Integration
      3. 10 scan average
      4. Wait 1 minute after each scan
      5. Stop after 2 hours
    5. Files saved as: Mb_OPD_H2O2_Citrate_H20_25_SCANNUMBER.txt
    6. 0.225mL Mb stock solution added just before the third scan
    7. This is denoted in the data file for the third scan
  • Note - This run was over in a hurry. Something was off (mass of Mb or mass of OPD or both) I stopped the run early and will use the balance in 207 for the remainder of experiments.

sample 2

  1. Stock solutions:
    1. Myoglobin
      1. 1.5mg (Mb:KCl lyophilized from Citrate, 5mg:500mg) in 1.0mL water
    2. OPD
      1. 3.0mg OPD in 3.3mL water
    3. H2O2
      1. 0.114mL 30% H2O2 in 0.887mL water
  2. Experimental Run
    1. Sample chamber set to 35C and to stir
    2. N2 run through the cuvette holder to reduce condensation on cuvette
    3. 2.025mL OPD stock solution
    4. 0.75mL H2O2 stock solution
      1. Start Data Collection
      2. 1ms Integration
      3. 10 scan average
      4. Wait 1 minute after each scan
      5. Stop after 2 hours
    5. Files saved as: Mb_OPD_H2O2_Citrate_H20_35_SCANNUMBER.txt
    6. 0.225mL Mb stock solution added just before the third scan
    7. This is denoted in the data file for the third scan

sample 3

  1. Stock solutions:
    1. Myoglobin
      1. Used the same stock as in sample 2
    2. OPD
      1. 2.5mg in 2.5mL
    3. H2O2
      1. 0.114mL 30% H2O2 in 0.887mL water
  2. Experimental Run
    1. Sample chamber set to 45C and to stir
    2. N2 run through the cuvette holder to reduce condensation on cuvette
    3. 2.025mL OPD stock solution
    4. 0.75mL H2O2 stock solution
      1. Start Data Collection
      2. 1ms Integration
      3. 10 scan average
      4. Wait 1 minute after each scan
      5. Stop after 2 hours
    5. Files saved as: Mb_OPD_H2O2_Citrate_H20_35_SCANNUMBER.txt Matt Hartings I messed up with the naming. Name should have 45 instead of 35. but file says 35
    6. 0.225mL Mb stock solution added just before the third scan
    7. This is denoted in the data file for the third scan



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