Investigation on how pH affects free and immobilised catalase enzymes.

Authors Avatar

Investigation on how pH affects free and immobilised catalase enzymes

Hypothesis

I hypothesis that as the ph of the solution that my enzymes are in (immobilised or free) moves away from neutral to more acidic or alkaline solutions that the amount of oxygen produced from the break down of hydrogen peroxide decreases. This can be explained by the research in which I stated how in acidic conditions enzymes are H+ acceptors and H+ donators in alkaline solutions this then breaks and destabilizes the ionic bonds denaturing the enzyme as it becomes unravelled. Also in my research I stated how the H+ ions would alter the globular shape of the protein distorting the active site. This distortion would affect the reaction rate as it would slow down due to the active site being distorted so it takes longer to break down the hydrogen peroxide and because more and more of the enzymes become distorted by the excess H+ or OH- ions meaning less active sites that are available to carry out the reaction. This can be proved by my preliminary test results, in which is showed when the ph was 13 or 2 the amount of oxygen produced was significantly reduced.

I finally hypothesis that the immobilised enzymes have a higher tolerance to ph this is due also to my research which showed that when immobilised enzymes were more tolerant to heat and so I can theorise that it may also be the same for changes in ph. This can come about by saying the beads in which the enzymes are in are protecting the enzymes. More concrete evidence for my hypothesis about how ph has less of an affect on immobilised enzymes are my preliminary test results on that show compared to the free enzymes they actually produce more oxygen, hence ph changes affect them less.

Introduction

Catalase occurs in many plant and animal tissue. It breaks down toxic Hydrogen peroxide, formed as a by-product of various biochemical reactions, into water and oxygen.

The activity of most enzymes is influenced by changes in pH. In this experiment, potato discs in solutions of known pH act on hydrogen peroxide, and the rate at which oxygen is evolved is measured. This reflects the activity of the Catalase in the potato.

Each enzyme has an optimal Ph, which is the Ph that it works the fastest and breaks the most substrates down in the least time. Most enzymes work best at a Ph of about   seven which is near neutral and our bloods natural Ph. They work the best at this temperature as they are proteins themselves and proteins can also become denatured by too acidic or alkaline conditions as well as high temperatures.  

Catalase the enzyme that I am using is a very important enzyme because it is present in every single cell in living organism. It works inside the cells of both animal and plants cells like in the liver for animals and potato cell for plants.  It breaks down hydrogen peroxide in the cells that it is present in; this makes it important, as hydrogen peroxide is a very poisonous substance produced by a lot of the chemical reactions in the body. Catalase ids found in large doses in the liver and the kidney which helps filter out waste products from the blood this makes it a suitable place to have a lot of catalase. Because a lot of hydrogen peroxide can be filtered and produced by the kidney and liver, which work continuously, they are also vital organs so catalase would be needed to break down hydrogen peroxide immediately before it can poison the organ that is disposing and filtering it.          

Enzymes

An enzyme is a biological catalyst. This is a substance that speeds up a reaction by lowering the activation energy, not getting used up and being able to be used many times. Enzymes speed up reactions by forming substrate complexes; pH is a factor that can affect the actual rate of reaction of an enzyme. Most enzymes are large globular proteins, which are very complex. They are two theories on how they break down substances the lock and key hypothesis and the induced fit theory. The induced fit theory is that as the enzyme and substrate approach each other it under goes a recognition process of conformational change in which the active site changes to fit the incoming substrate. And as the substrate binds to the active site creating a substrate enzyme complex the binding process stretches or compresses one or more of the chemical bonds in the substrate this is like a kind of twisting affecting which explains how it is so that enzymes reduce the activation energy.

The lock and key theory is simply how the enzyme has a site that already has the perfect shape for the substrate and so when they connect and meet the substrate enters the active site and is broken down. There are two main ways in which ph can affect an enzyme and they are.

The amphoteric affect

Enzymes are tertiary and quaternary proteins meaning that they have been folded over to for a three dimensional shape. This folding of the protein is enabled and strengthened by ionic bonds, which is the static attraction between proteins. It is this properties that makes proteins buffers as they can act as H+ acceptors or donators this then helps to keep the surroundings more neutral conditions. This is why pH affects enzymes as in extreme acidic conditions the amine end of the folded protein takes up a H+ ion to become NH3+ and in extreme alkaline conditions the carboxylic group loses a H+ ion to become COO-. It is this actual fact that can cause denaturing of proteins destabilising the ionic bonds affecting the active site which is the part of the enzyme that carries out the reaction resulting in the enzyme no longer being able to carry out its purpose.  

Join now!

Non active site inhibition

This is simply the fact that inorganic ions like Na or H can distort the shape of enzymes by binding to allosteric cites which are a kind of activation site away from the main active site thus changing the globular structure and in turn the shape of the active site so no substrate may enter or fit.

Another type of non active site directed inhibition is when an ion just simply binds to a part of the enzyme and changes its shape so it is denatured when there is no active site.

...

This is a preview of the whole essay