Vernier Software & Technology

# Analysis of Enzymes using Tyrosinase

## Introduction

Enzymes are molecules that regulate the chemical reactions that occur in all living organisms. Almost all enzymes are globular proteins that act as catalysts, substances that speed up chemical reactions. Enzymes catalyze reactions by reducing the activation energy for a specific reaction to occur and yet are neither destroyed nor altered during this process. At the molecular level, enzymes catalyze these reactions by briefly binding to the substrate or reactants to form an enzyme-substrate complex. The reaction takes place while the substrate is bound to the enzyme, converting the substrate to the new product. The new product is then released from the enzyme substrate complex and the enzyme is then free to bind with more substrate.

This investigation is designed to introduce you to the quantitative analysis of enzymes using the enzyme tyrosinase. Tyrosinase is an enzyme that is involved in melanin synthesis, which gives skin its color. Tyrosinase is also involved in the browning of fruits, tubers, and fungi that have been damaged.In the presence of oxygen (O2), tyrosinase (E) catalyzes the hydroxylation of tyrosine into the colorless compound 3,4-dihydroxyphenylalanine, or DOPA for short. Tyrosinase then catalyzes DOPA into dopaquinone which spontaneously converts into dopachrome. Dopachrome will eventually be turned into melanin. Dopachrome is an orange colored compound with a peak absorbance at 475 nm that can be monitored using a spectrophotometer.

The first step in this series of reactions is the rate-limiting step for the entire process. This means that tyrosinase is much better at converting DOPA to dopaquinone than it is at converting tyrosine to DOPA. Knowing which step is the slowest in a complex biochemical process can be medically important. People with Parkinson’s disease have low levels of dopamine in their brain. Dopamine is synthesized from tyrosine; the first step involves converting tyrosine to DOPA, which is also the rate-limiting step for the entire process. As a result, DOPA is given to people with Parkinson’s disease to increase their natural production of dopamine. This, in turn, alleviates the symptoms of the disease.

## Objectives

In this Preliminary Activity, you will use a spectrophotometer to measure color changes due to the production of dopachrome and determine rate of reaction.

After completing the Preliminary Activity, you will first use reference sources to find out more about enzymes, tyrosine, and tyrosinase before you choose and investigate a researchable question. Some topics to consider in your reference search are:

• catalyst
• enzyme
• substrate
• tyrosine
• tyrosinase
• DOPA
• Michaelis-Menten constant
• Lineweaver-Burk plot

## Sensors and Equipment

This investigation features the following Vernier sensors and equipment.

You may also need an interface and software for data collection. What do I need for data collection?

## Investigating Biology through Inquiry

See other experiments from the lab book.

 1 Buffer Investigations 2 Diffusion 3 Investigating Osmosis 4 Chemistry of Membranes 5 Investigating Protein 6A Testing Catalase Activity (O2 Gas Sensor) 6B Testing Catalase Activity (Gas Pressure) 6C Testing Enzyme Activity (Spectrometer) 7 Introduction to Biofuels: Enzyme Action 8 Analysis of Enzymes using Tyrosinase 9 Cellular Respiration 10A Sugar Metabolism with Yeast (Carbon Dioxide Gas) 10B Sugar Metabolism with Yeast (Ethanol) 11 Fermentation with Yeast 12 Photosynthesis by Chloroplasts 13 Transpiration of Plants 14 Plant Pigments 15 Heart Rate 16 Investigating Dissolved Oxygen 17 Investigating Primary Productivity 18 Modeling Population Dynamics 19 Water Monitoring 20 Evolution of Cellobiase in Fungi 21 Introduction to Molecular Evolution 22 Evolution of Yeast

### Investigation 8 from Investigating Biology through Inquiry Lab Book

#### Included in the Lab Book

Vernier lab books include word-processing files of the student instructions, essential teacher information, suggested answers, sample data and graphs, and more.