{"id":2515,"date":"2021-09-15T13:51:34","date_gmt":"2021-09-15T17:51:34","guid":{"rendered":"https:\/\/wordpress-projects.wolfware.ncsu.edu\/bio-183-lab-yhnn8gz\/?page_id=2515"},"modified":"2023-05-10T21:24:36","modified_gmt":"2023-05-11T01:24:36","slug":"unit-2-labs-2-3-enzyme-lab-make-up-module","status":"publish","type":"page","link":"https:\/\/wordpress-projects.wolfware.ncsu.edu\/bio-183-lab-yhnn8gz\/unit-2-labs-2-3-enzyme-lab-make-up-module\/","title":{"rendered":"Unit 2: Labs 2-3 Enzyme Lab \u2013 Make Up Module"},"content":{"rendered":"\n

Enzyme Lab Handout (covers 2 weeks of Online Make Up labs)<\/a> \u2013  Includes areas for filling out your tables and answering questions.<\/p>\n\n\n\n

Make sure you have Excel or access to Google Sheets.  As an NCSU student, you can download Microsoft Office 365 from NC State\u2019s OIT<\/a> software site for free!<\/p>\n\n\n\n

Read along in your Enzyme Handout as you work with the information in this webpage.  Fill in and answer questions accordingly.  Your Enzyme Handout will be collected and graded later.<\/em><\/h4>\n\n\n\n
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\"Enzyme<\/figure><\/div><\/div>\n\n\n\n
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\"Effect<\/figure><\/div><\/div>\n<\/div>\n\n\n\n

How Cells Work \u2013 Enzymes<\/a><\/h3>\n\n\n\n

Chemical Reactions and Activation Energy<\/h3>\n\n\n\n
\"Graph<\/figure>\n\n\n\n

Catalase<\/span><\/strong><\/p>\n\n\n\n

\"Catalase
Catalase<\/figcaption><\/figure>\n\n\n\n

Animation \u2013 Enzyme Function and the difference between competitive and non-competitive inhibitors.<\/strong><\/a><\/p>\n\n\n\n

<\/div>\n\n\n\n

Activity 1: Qualitative Study of Catalase<\/h3>\n\n\n\n

2H2<\/sub>O2<\/sub> + CATALASE \u2014-> 2H2<\/sub>O + O2<\/sub> + CATALASE<\/h3>\n\n\n\n

Catalase is prepared by blending a potato in cold water, then filtered by straining the mixture through 4 layers of cheesecloth.  The catalase solution is then kept cold on ice.  Why would filtering and maintaining a cold temperature be important for the catalase solution?<\/p>\n\n\n\n

\"\"<\/figure>\n\n\n\n

Procedure A Testing the Activity of Catalase<\/h3>\n\n\n\n

Follow steps 1-3 and complete Table 1 using the results in the pictures below.  The picture on the left shows the appearance of catalase and boiled catalase.  What would account for the difference in appearance of the boiled catalase?  Where did the \u201cchunks\u201d come from?  The picture on the right shows the reaction in tubes 1, 2, 3, and 4 after 5 minutes of the catalase reaction.<\/p>\n\n\n\n

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Procedure B Testing the Activity of Catalase Under Six Different Environmental Conditions<\/h3>\n\n\n\n

You will be assigned to work with ONE of the six environmental conditions (Groups) below.  Propose a plausible hypothesis for your assigned experiment.  The same basic reaction as tube #3 in Procedure A is followed, but treated with an assigned environmental condition.  Answer all questions and fill in the Table for your assigned treatment in your lab manual.   <\/strong><\/p>\n\n\n\n

Your assigned Group for Procedure B will be the same Group assignments for Procedure D (further below). <\/strong><\/p>\n\n\n\n

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\nGroup I: Effect of Substrate Concentration on Catalase<\/figcaption><\/figure>\n<\/div>\n\n\n\n
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Group II: Effect of Hydroxylamine and Subs. Concentration<\/figcaption><\/figure>\n<\/div>\n<\/div>\n\n\n\n
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Group III: Effect of Salt Concentration on Catalase<\/figcaption><\/figure>\n<\/div>\n\n\n\n
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Group IV: Effect of the Metal Copper Sulfate<\/figcaption><\/figure>\n<\/div>\n<\/div>\n\n\n\n
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Group V: Effect of pH*<\/figcaption><\/figure>\n<\/div>\n\n\n\n
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Group VI: Effect of Temperature<\/figcaption><\/figure>\n<\/div>\n<\/div>\n\n\n\n
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pH Test strip reading<\/figcaption><\/figure>\n\n\n\n

<\/p>\n<\/div>\n\n\n\n

<\/div>\n<\/div>\n\n\n\n

Activity 2: Quantitative Study of Catalase<\/h3>\n\n\n\n

2H2<\/sub>O2<\/sub> + CATALASE + GUAIACOL (COLORLESS) \u2014-> 2H2<\/sub>O + CATALASE + TETRAGUAIACOL (ORANGE-BROWN)<\/h3>\n\n\n\n

We can study the same reaction as above in a more detailed manner by fine tuning the experiment.  Instead of learning about the relative activity of catalase, observing more or less reaction, we can introduce a sensitive color indicator to detect the presence of one of the reaction products.  The color indicator guaiacol will change from clear to orange-brown as it binds to the O2<\/sub> produced in this reaction forming tetraguaiacol.  This color change can be measured using a spectrophotometer.  These data can then be graphed and rates determined for the baseline reaction of catalase, but also for the various treatments.<\/p>\n\n\n\n

Procedure C Using a Spectrophotometer to Monitor Baseline Catalase Activity<\/h3>\n\n\n\n

Carefully read the Procedure C background.  This will help you fill in and answer the questions and Table 9 in within the Procedure C directions in your lab handout.  We will use a Genesys spectrophotometers and square disposable cuvettes.  If you have not used a spectrophotometer before, view the videos below.  It is very important to use an appropriate blank for each treatment that is run.  The blank will contain all the same components as the treatment tube, minus the enzyme.  This way we will know that any change in color is resulting from the activity of the enzyme itself.<\/p>\n\n\n\n

How spectrophotometers work<\/a> \u2013 transmittance and absorbance<\/h3>\n\n\n\n

How to use a Genesys 10 Spectrophotometer<\/a><\/h3>\n\n\n\n

Sample Data for Procedure C<\/strong><\/a><\/p>\n\n\n\n

<\/p>\n\n\n\n

Use this data to create a graph monitoring the baseline catalase reaction.  Be sure to label the graph correctly with all appropriate graphing components outlined by your TA.<\/p>\n\n\n\n

Determining  Baseline data<\/strong> –  Be sure to label the graph correctly with all appropriate graphing components outlined by your TA and indicated in the Resources \u2013> Appendix D: Graphing Using Excel.<\/p>\n\n\n\n