Can you discover the conditions that affect the rate of enzyme catalyzed reactions? …And make them glow!
This lab allows students to investigate enzyme activity and the conditions that affect it. Under optimal conditions enzymes catalyze, or speed up the rate of, chemical reactions. In this lab, a substrate that is not fluorescent under basal conditions, will fluoresce under blue light when sugar molecules are removed through hydrolysis by the enzyme beta-galactosidase. Observing how brightly the solution fluoresces over time allows students to observe the reaction progress, and to quantify the effect of different factors on enzyme efficiency.
This Lab is structured as four inquiry-based investigations. In each investigation, students explore the effect of a different factor on enzyme reaction rate:
- enzyme and substrate concentration
- competitive inhibition.
This lab provides enough reagents for eight lab groups to each pursue at least two of these investigations (32 students):
- Substrate for enzymatic reaction
- Purified Beta-Galactosidase enzyme solution
- Dilution buffers (2)
- Inhibitor solutions (2)
- Alkaline medium
- Fluorescent solution for standard curve
- This activity is compatible with blueGel™, blueBox, P51™, or other blue light transilluminator systems
Required equipment (one per lab group):
- Blue light transilluminator (such as blueBox™ or blueGel™) or P51™ Molecular Viewer
- 2-20 ul micropipette
- Suggested for temperature investigations: miniPCR™ thermal cycler (ideal set up) or water bath and floaties, iced water.
- Techniques utilized: Micropipetting, fluorescence detection of enzyme activity.
- Time required: can be completed in one 45 min. class period, or extended into two 45-min periods or a single block.
- Reagents needed: P51 Enzyme Lab: β-Gal Glow™ (this product).
- Suggested skill level: Familiarity with protein structure and enzyme activity, from middle school to college.
- These reagents require refrigerated storage.
- Shelf-life: 6 months when stored at 4 degrees.
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