Lab 4: Gen Bio Lab

Secondary Structure

*Hydrogen Bonds* form between the backbone of adjacent amino acids Common secondary structures: *alpha-helix* and *beta-sheet*

Enzyme Assay

the rate of product formation is proportional to the amount of enzyme present Can measure the *accumulation of products* . or . the *disappearance of substrate*

*Denaturation*

disruption of the enzyme's shape by breaking the weak bonds that maintain the native shape

Substrate

A specific reactant acted upon by an enzyme

Experiment 2: Inhibiting the Action of Catechol Oxidase

*Phenylthiourea* (PTU) is an enzyme inhibitor. Setup: Looked at 3 test tubes with potato extract and cathecol. - one control with no PTU Seeing if competitive or noncompetitive by looking at color in solution. - The darker we saw the more reactive was the . enzyme. Results: We found it is competitive because the more catechol we added, the darker the brown color showing that its competitive because increased concentration of Catechol increased brown (result), if it was noncompetitive the increase would not have mattered

Experiment 1: Experimental Method and the Action of Catechol Oxidase

*Process:* Substrate: Catechol Product: Benzoquinone (appears brown) Enzyme: Potato Extract Many fruits and veggies turn brown in presence of oxygen due to catechol Combine catechol w/ potato extract and look for enzyme activity (brown color) We just added these together in 3 tubes and checked to see if color changed. *Dependent*: Color (enzyme activity) *Independent*: concentration of catechol and potato extract *Control*: There were 2 control tubes that did not combine the catechol and the potato extract that did not turn color *Results*: Color changed when potato extract and enzyme together

Activity of Enzymes Influenced by:

*Temperature*: high temperature denatures enzyme, low temperature inhibits enzyme *pH*, *osmotic pressure* etc.

Enzyme Characteristics

- Speed up the rate of reaction - Are not used up in the process - Are usually proteins - Are veery substrate specific (active site)

Experiment 3: Influence of Concentration, pH, and Temperature on the Activity of Amaylase

1. Figure out Enzyme concentration to use (different percentages of amaylse and time the reaction using indicator for starch, so darker meant more starch, less reaction) - using I2KI pick one that you see it doesn't get used up too fast so we can see results - Turns from dark to light color as it disappears 2. Test pH variance of that concentration - in puffers pH 4, 5, 6, 7, 8, 9 pH is ideal at 6.5-7 which is the mouth pH where amylase thrives, so light color - protein denatures in too low or two high pH and we see starch stay so dark color 3. Test temperature variance of that concentration - do in temperatures 80C, 37C, room temp, (-22C) - We found it was ideal at 37 (lightest) thats body temp. - Temperature protein denatures at high so stays super dark - Less effective at low temperatures so stays pretty dark in I2Ki *Dependent*: Breakdown of Starch to glucose *Independent*: Temp, PH, concentration *Control*: ?? Testing Factors that Affect amylase activity: 1. Enzyme concentration 2. pH (best pH is 6/7, fastest so darker at this) 3. Temperature ( less starch lighter, more starch darker)

Protein's 3D Structure

1. Primary Structure: linear order of amino acids 2. Secondary Structure: localized folding into pleated sheets (beta) or (alpha) 3. Tertiary Structure: 3-D Shape of the entire protein 4. Quaternary: Combination of more than one polypeptide .

Endergonic

Absorb energy products store more energy than reactants non spontaneous

Metabolism

All the chemical reactions in the body Food provides nutrients, nutrients get converted to building blocks or energy in our body through chemical reactions

Enzymes

Catalysts for chemical reactions in living things. Effective catalysts because they reduce the activation energy for reactions.

Activation Energy

Energy needed to get a reaction started Even energetically favorable reactions require activation energy.

Noncompetitive Inhibition

Enzyme has two binding sites: - Active Site - Regulatory Site Inhibitor binds to the regulatory site, changing the conformation of the enzyme - Slows down enzyme activity - cannot be reversed by increased concentrations of substrate

The role of melanin in animals, especially in human skin where it helps protect against harmful ultraviolet rays, is relatively well supported; however its role in plants is inconclusive. Speculate about a possible role for catechol melanin in plant tissue?

In plants, catechol oxidase plays a key role in enzymatic browning by catalyzing the oxidation of catechol which can rapidly form the melanin that grants damaged fruits their dark brown coloration. ??

Amaylase

Is a product in salivary glands and in pancreas Breaks down amylose in starch into maltose

Metabolic Pathway

Many chemical reactions happen sequentially, with the product of one reaction serving as a reactant for the next reaction. Each reaction is catalyzed by a specific enzyme . Most enzymes are proteins, and the molecules they bind in the reaction are called substrates

Peptide Chains

Proteins are made of *amino acids* Amino acids are linked by *peptide bonds* formed between the carboxyl group of one amino acid with the amino group of another Each polypeptide chain has directionality: *N-terminus* to *C-terminus*

Exergonic

Release energy Reactants store more energy than products spontaneous

An exergonic (spontaneous) reaction is a chemical reaction that __________.

Releases energy when proceeding in the forward direction

Catechol

Substrate that turns brown when reacted with potato extract. Product is brown

Anabolism

Synthesis fo large macromolecules in the cell - Requires input of energy

Competitive Inhibition

The inhibitor resembles the enzyme's normal substrate. - mimic prevents the actual substrate from binding Can be reversed if the concentration of the substrate is raised to sufficiently high levels

Tertiary and Quaternary Structures

Various interactions between *R groups* of amino acids

active site

a region on an enzyme that binds to a protein or other substance during a reaction. Where the substrate binds

Enzymes can be used to accelerate anabolic or catabolic reactions?

both

Catabolism

breaking down of large molecules into smaller molecules - Release of energy