Name_____________________________ Date____________Building A Protein: simulating TRANSLATION

Summary

This hands-on activity is designed to simulate how proteins are assembled by ribosomes (this process is called translation). The protein that you will translate is ADH (antidiuretic hormone). In this activity you will assemble the amino acids that are the building blocks for this protein. You will then simulate how the ribosome “reads” the sequence of amino acids from a strand of mRNA. Finally, you will create the peptide bonds that join the amino acid monomers together and the disuphide bonds that give proteins their globular shape (see your class notes). Your final product should have eight peptide bonds, creating one chain, and eight molecules of water. DISCLAIMER: In reality, the strand of mRNA has “codons” for each amino acid. I’ve left this part out for now for simplicity. However, the start and stop “codons” are included to give you a sneak peak of what’s to come.

Learning Goals

· Describe the basic structure of amino acids.

· Explain how peptide bonds are formed.

· Describe how the chemical nature of amino acid side chains (R group) determine the shape and tertiary structure of the protein.

· Define translation.

Materials:

· Amino Acid Template worksheet (one per student)

· Paper stand of “mRNA”

· 1 Scissor, and 1 roll of tape

Preparation for the activity:

1. Assemble the strand of mRNA.

Cut along the solid lines and tape the three strips of paper together. The second strip that begins with "glu" should be taped to strip number one that ends in "phe". Then tape strip number three that begins with "gly" to strip number two that ends with "arg". You should have one strip of paper with eleven mRNA codons.

2. Assemble the amino acids.

Cut along the solid lines on the Specific Amino Acid worksheet. You will have nine amino acid cards. Then cut out the nine R-groups. Tape one amino acid to one R-group card to create nine specific amino acids.

3. Cut out the nine bonds and the eight process labels.

Procedure:

1. Locate the start “code” (called a codon) on the mRNA strand, this tells the ribosome to attach and begin. Then using your amino acid table, find the cys amino acid and tape it onto the mRNA strand.

2. Find the next amino acid instructed by the mRNA and tape it into place.

3. “Perform” dehydration synthesis to link the two amino acids together in the following way.

a. Cut out the -OH of the carboxyl group on the first amino acid.

b. Cut out the -H of the amino group on the second amino acid.

c. Combine these two piece (-OH and -H ) to form a water molecule.

d. Tape a peptide bond label between the carbon of the first amino acid and the nitrogen of the second amino acid.

e. Lay the dehydration synthesis label below the peptide bond, and above the water molecule.

4. Repeat this procedure until you reach the stop “code”, UAA. Your primary structure is complete.

5. Now tape the disulphide bond between the two cystine amino acids. This is referred to as a disulphide bridge, and it is ONE of the bond types that hold a protein chain in its tertiary structure. Note this simulation ignored the hydrogen bonds that create the secondary structure (see your class notes, you will need to know this).

THE STAGES OF PROTEIN FORMATION:

The sequence of amino acids is called the primary structure.

(Remember, it is at the ribosome where the free floating amino acids are assembled into their primary structure)

Then hydrogen bonding between the amino acids causes the polypeptide to fold and twist forming the secondary structure. There are two main types of secondary structures, the alpha helix or beta sheets.

Note: it is the repeating part of the chain that is interacting, this is why ALL proteins have the same secondary folding pattern.

Tertiary structure:

The alpha helixes and beta sheets are then folded into a compact globular structure. A protein’s tertiary structure is unique because it is determined by the “R” group of each amino acid.

Quaternary structure:

If another protein, chemically bonds to another. The resulting molecule takes the form of the tertiary structure.

ENZYMES:

https://www.youtube.com/watch?v=XTUm-75-PL4

Enzymes are biological molecules (proteins) that significantly speed up the rate of virtually all of the chemical reactions that take place within cells.

They are vital for life and serve a wide range of important functions in the body, such as aiding in digestion and metabolism.

· Some enzymes help break large molecules into smaller pieces, catabolism.

· Other enzymes help bind two molecules together to produce a new molecule, anabolism.

· Enzymes are highly selective catalysts, meaning that each enzyme only speeds up a specific reaction. The binding site on the enzyme is substrate specific.

· The molecules that an enzyme works on are called substrates.

· During these chemical reactions the enzyme itself is not used up or chemically changed. Enzymes are recycled and just a few can catalyze many reactions.

HOW ENZYMES CATALYZE REACTIONS: THE INDUCED FIT MODEL

ENZYMES: LOWER THE ACTIVATION ENERGY REQUIRED FOR REACTIONS:

To see this in action go to PhET Simulations: Reactions & Rates

ENZYMES OFTEN WORK TOGETHER IN A CHAIN OF REACTIONS CALLED A “METABOLIC PATHWAY”. The product of the first enzyme becomes the substrate for the second.

REGULATING ENZYME ACTION:

In order to understand how enzymes are controlled we can look at how a thermostat works.

In order to maintain a constant internal environment there needs to be signals that turn the thermostat on and off. The enzymes in our body must also be able to turn on and off using chemical signals.

Enzyme activity can be regulated in several ways:

In biochemistry, allosteric regulation (or allosteric control) is the regulation of a protein by binding an effector molecule at a site other than the protein's active site. Effectors that enhance the protein's activity are referred to as allosteric activators, whereas those that decrease the protein's activity are called allosteric inhibitors.

Allosteric inhibition (as shown below) the bonding of the inhibitor to the allosteric site changes the shape of the active sight. As a result, the substrate is not able to bind as tightly and the enzyme’s activity slows down. REMEMBER: the active site is SPECIFICALLY shaped to fit one type of substrate.

Allosteric activation (b) in this case, the binding of the activator to the allosteric site causes the active site to obtain the correct shape and the enzymes attraction to its substrate is enhanced.

NOTE: The allosteric enzyme is still functional with or without the the affector (see graph above), the affector simply enhances and inhibits the activity.

Sometimes the LAST product of a metabolic pathway (series of reactions) acts as an allosteric inhibitor to one of the first enzyme along the chain. This is called negative feedback

If the LAST product of a metabolic pathway acts as an allosteric activator to one of the first enzymes along the path, we call this positive feedback.

COENZYMES & COFACTORS: Substances that are REQUIRED for the enzyme to function.

· Metal ions are usually cofactors.

· Coenzymes are are organic molecules that are often, derived from vitamins. 

This is just one of the reasons it is so important to eat a well balanced diet.

Homeostatic Imbalance – Vitamin C Deficiency

E.g. Scurvy - “The Curse of the Arctic” Early Arctic explorers had a diet which lacked fresh meat, fruit and vegetables for years at a time. As a result they suffered from scurvy - a disease caused by the lack of Vitamin C. Vitamin C is a cofactor necessary to produce the protein collagen. Collagen is a major component of connective tissue. Without Vitamin C, a person’s connective tissues will break down and they will eventually die. The symptoms of scurvy are brutal - blood vessels rupture and blood weeps from hair follicles. Blotches appear all over the body and any bruise or exertion could result in internal bleeding. Arms and legs swell, and wounds, even small scratches don’t heal. Skin becomes dough-like and the gums red, and blackened until teeth become so loose they fall out. Slowly, the body’s tissues fall apart as their collagen scaffolding disintegrates.

CLASSIFICATION OF ENZYMES BY REACTION TYPE: most names end with -ase

ASSIGNMENT: FACTORS AFFECTING THE RATE OF ENZYME ACTIVITY

Using the internet or other textbook resources explain how each of the following affects the rate of enzyme activity. INCLUDE A GRAPH whenever possible.

1. pH

2. temperature

3. substrate concentration

4. enzyme concentration

Be prepared to share your answers.