Sandbox2O14: Difference between revisions

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== Figure 4 ==

Protein test using 10µL of protein from elution and 30mg of PNPP and 3mL of PNPP solution. Measuring absorbance at 405M.

== Figure 5 ==

Protein test using 10µL of protein from elution 2 and 30mg of PNPP and 3mL of PNPP solution. Measuring absorbance at 405M.

== Figure 6 ==

Protein test using 10µL of protein from elution 1 and 30mg of PNPP and 3mL of PNPP solution, changing the pH to about 5.60. Measuring absorbance at 405M.

== Figure 7 ==

Image of SDS-PAGE gel that proves that protein was present in the sample.

The following week two more experiments were run, one with a 1:1 ratio of PNPP and PNPP solution with 10µL of protein solution for elution 1. The first was just with the PNPP mixture and showed no positive results, with absorbance values bouncing back and forth between values in the range of 0.15 to 0.096. After this test, a drop of 3.0M HCL was added to the solution to increase acidity slightly, which harbored almost identical results to the previous test. After, an experiment with PNPA was performed to see if a change in substrate would cause the protein to react. Figures 8, 9, and 10 show the results of these reactions.

== Figure 8 ==

Protein test using 10µL of protein from elution 1 and 3mg of PNPP and 3mL of PNPP solution. Measuring absorbance at 405M.

== Figure 9 ==

Protein test using 10µL of protein from elution 1 and 3mg of PNPP and 3mL of PNPP solution and 1 drop of HCL with a pH of about 10.50. Measuring absorbance at 405M.

== Figure 10 ==

Protein test using 10µL of protein from elution 1 and 10mg of PNPA and 10mL of PNPP solution and 1 drop of HCL with a pH of about 10.50. Measuring absorbance at 405M.

The following week two more reactions were run to prove that the protein function was as an acetyl esterase. The second to last experiment was run with a ratio for 5:1 PNPP and PNPP solution which resulted in no absorbance change throughout the experiment. Then as a final used PNPA with a 5:1 ratio solution, and during this experiment 5µL of protein solution was added to the cuvette solution being measured. This was done to see if more protein concentration was needed to perform the reaction, and after 30 minutes and 155µL of protein solution, no absorbance change was observed. Figures 11 and 12 show these reactions.

== Figure 11 ==

Protein test using 10µL of protein from elution 1 and 15mg of PNPA and 3mL of PNPP solution. Measuring absorbance at 405M.

== Figure 12 ==

Protein test with increasing protein solution of 5µL every minute from elution 1 with 15mg of PNPA and 3mL PNPP solution. The pH of the solution is 8.35 and it was measuring absorbance at 405M.

This data shows two possible case results. One is that the protein sample might have been out of a cold environment for too long which would cause the protein to denature and not operate as predicted. Another potential case is that the predicted function of the protein was incorrect.

</StructureSection>

== Figure 1 ==

Lowest Calculated affinity image from SwissDock of -6.133 kcal/mol, with the ligand 2-acetamido-2-deoxy-beta-D-glucopyranose.

== Figure 2 ==

The second lowest calculated binding affinity value, -5.629 kcal/mol, predicted bonding site. Believed that the protein’s binding site is in the center between the beta sheet group on top and the alpha helices on the bottom with the ligand 2-acetamido-2-deoxy-beta-D-glucopyranose.

Sandbox2O14: Difference between revisions

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@@ Line 47: / Line 47: @@
 == Figure 4 ==
 Protein test using 10µL of protein from elution  and 30mg of PNPP and 3mL of PNPP solution. Measuring absorbance at 405M.
+== Figure 5 ==
+Protein test using 10µL of protein from elution 2 and 30mg of PNPP and 3mL of PNPP solution. Measuring absorbance at 405M.
+== Figure 6 ==
+Protein test using 10µL of protein from elution 1 and 30mg of PNPP and 3mL of PNPP solution, changing the pH to about 5.60. Measuring absorbance at 405M.
+== Figure 7 ==
+Image of SDS-PAGE gel that proves that protein was present in the sample.
+The following week two more experiments were run, one with a 1:1 ratio of PNPP and PNPP solution with 10µL of protein solution for elution 1. The first was just with the PNPP mixture and showed no positive results, with absorbance values bouncing back and forth between values in the range of 0.15 to 0.096. After this test, a drop of 3.0M HCL was added to the solution to increase acidity slightly, which harbored almost identical results to the previous test. After, an experiment with PNPA was performed to see if a change in substrate would cause the protein to react. Figures 8, 9, and 10 show the results of these reactions.
+== Figure 8 ==
+Protein test using 10µL of protein from elution 1 and 3mg of PNPP and 3mL of PNPP solution. Measuring absorbance at 405M.
+== Figure 9 ==
+Protein test using 10µL of protein from elution 1 and 3mg of PNPP and 3mL of PNPP solution and 1 drop of HCL with a pH of about 10.50. Measuring absorbance at 405M.
+== Figure 10 ==
+Protein test using 10µL of protein from elution 1 and 10mg of PNPA and 10mL of PNPP solution and 1 drop of HCL with a pH of about 10.50. Measuring absorbance at 405M.
+The following week two more reactions were run to prove that the protein function was as an acetyl esterase. The second to last experiment was run with a ratio for 5:1 PNPP and PNPP solution which resulted in no absorbance change throughout the experiment. Then as a final used PNPA with a 5:1 ratio solution, and during this experiment 5µL of protein solution was added to the cuvette solution being measured. This was done to see if more protein concentration was needed to perform the reaction, and after 30 minutes and 155µL of protein solution, no absorbance change was observed. Figures 11 and 12 show these reactions.
+== Figure 11 ==
+Protein test using 10µL of protein from elution 1 and 15mg of PNPA and 3mL of PNPP solution. Measuring absorbance at 405M.
+== Figure 12 ==
+Protein test with increasing protein solution of 5µL every minute from elution 1 with 15mg of PNPA and 3mL PNPP solution. The pH of the solution is 8.35 and it was measuring absorbance at 405M.
+This data shows two possible case results. One is that the protein sample might have been out of a cold environment for too long which would cause the protein to denature and not operate as predicted. Another potential case is that the predicted function of the protein was incorrect.
+</StructureSection>
+== Figure 1 ==
+Lowest Calculated affinity image from SwissDock of -6.133 kcal/mol, with the ligand 2-acetamido-2-deoxy-beta-D-glucopyranose.
+== Figure 2 ==
+The second lowest calculated binding affinity value, -5.629 kcal/mol, predicted bonding site. Believed that the protein’s binding site is in the center between the beta sheet group on top and the alpha helices on the bottom with the ligand 2-acetamido-2-deoxy-beta-D-glucopyranose.