This post is just to share a recent experiment I did but before we get to that lets just take a minute and look at this logo.
To my knowledge this logo is misleading firstly the mixture they have is not 100% pure protein as the side of the box says it contains 15mg sodium in addition to 6g of proteins but I’m not here to argue that look at the back of the box.
It says , “Indulge gelatine contains important proteins for healthier skin, hair and nails.” This is misleading since we all know that gelatine is a hydrolysed form of the protein collagen and even though collagen does make up our skin and connective tissue we know that it is keratin that makes up our hair and nails and karetain and collagen are 2 completely different proteins.
- Collagen is made of 3 peptide chains which are left handed helix. These 3 helices are then coiled into a right handed coil (triple helix) forming the collagen structure. The peptide chains are predominantly made of glycine and proline and they both serve special roles within the molecule. Glycine works to stabilize since it is the smallest amino acid it fits on every third position of the chains which end up in the interior of the collogen structure or the center of the triple helix. Proline because of its cyclic structure aids in the formation of the coils in the triple helix structure of the collagen, since proline changes the direction of the chain due to is lack of maximum h bonds that can be formed.
- Keratin is a fibrous structural protein of hair, nails, horn, hoofs, wool, feathers, and of the epithelial cells in the outermost layers of the skin. The polypeptide chains of keratin are arranged in parallel sheets held together by hydrogen bonding.Of the amino acids in keratin, cystine may account for as much as 24 percent. The numerous disulfide bonds formed by cystine are responsible for the great stability of keratin: it is completely insoluble in hot or cold water and is not attacked by proteolytic enzymes (the enzymes that cleave protein molecules).
Gelatine is made from collagen thus according to the research it is rich in Proline and glycine both of which are non essential amino acids meaning our body makes them we do not get them from our diet….. thus the box is misleading, Further more it says that the gelatine is important for hair and nails and since we know that keratin is responsible for hair and nails and keratin is mainly made of cystine molecules the box is again wrong. (If it was made of cystine and keratin the following experiment would not have work as stated proteolytic enzymes such as papain doesn’t work in it)
Now the video which i made based on the enzyme papain and how it works (its my first video so please feel free to give any tips)…..
Km – the rate constant or it can be explained as how much substrate concentrationis required by an enzyme to reach to the half of maximum rate or velocity of enzyme.
Vmax- that point is the saturation point or maximum substrate concentration to have maximum rate of the reaction.
So what are the effects of changing variables on enzymic reactions?
What if we increased the substrate concentration…
As the concentration of substrate increases, the rate of reaction also increases until the point saturation occurs. It means as you increase the concentration, rate keeps increasing and then one point comes when all the enzymes are occupied and there is excess substrate. Thus after this point, increasing the concentration will result in the graph tapering off.
Effect of temperature on enzyme activity
If the temperature is too high, there is enough energy to break down the tertiary structure of the enzyme this alters the shape of the active site so that the substrate no longer fits; the enzyme is said to be denatured. However if the temperature is to low the substrate molecules simple do not have enough energy and so the reaction cannot take place.
Effect of pH on enzyme activity:
The optimum pH for each enzyme is different based on the job of the enzyme and where they work. Some enzymes work in acidic conditions such as those in our stomach like pepsin while others work under basic or normal conditions. This is because of the proteins that might be present at the active site and just like the effect of temperature deviation for the optimum pH can cause a conformational change in the shape of the active site.
OH INHIBITORS !!!!! : (
Inhibitors are molecules that combine to enzymes and prevent the substrate from attaching to the active site by changing the shape of the active site or the affinity of the enzyme these result in decreses in the rate of reactions which can be plotted on a graph called the Linweaver – Burk plot or even the Michealist- menten curve.
There are many types of inhibitors some are reversible while others are irreversible:
Some reversible inhibitors are:
Competitive inhibition (Slide B)
Substrate and inhibitor cannot bind to the enzyme at the same time, the inhibitor having an affinity for the active site of an enzyme and the inhibitor having a complementary shape that resembles the substrate. Thus inhibitor compete for access to the enzyme’s active site. This type of inhibition can be reversed by increasing the concentrations of substrate thus the vmax is left unchanged however since there is more substrate the Km or 1/2Vmax point will be increased.
Non-competitive inhibition(Slide C)
Bind at different sites than the active site. When both the substrate and the inhibitor are bound, the enzyme-substrate-inhibitor complex cannot form product and can only be converted back to the enzyme-substrate complex or the enzyme-inhibitor complex. The Vmax is decrease since the enzyme substrate complex or the active state is disrupted, however the Km remains the same since binding of the enzyme to the substrate is not affected.
Uncompetitive inhibitor (Slide D)
Enzyme inhibitor binds only to the complex formed between the enzyme and the substrate the Vmax decreases as well as the Km at equal proportions since formation of the enzymes substrate complex is affected or however the enzyme affinity is increased.
The inhibitor may bind to the enzyme whether or not the enzyme has already bound the substrate but has a greater affinity for one state or the other. The binding of the inhibitor to the enzyme reduces its activity but does not affect the binding of substrate. Vmax will decrease due to the inability for the reaction to proceed as efficiently, but Km will remain the same as the actual binding of the substrate, by definition, will still function properly.
Learning about the different types of inhibitors was new to me and was sort of challenging but once you understand the basics of the Lineweaver- burk plot and how each inhibitor works you can easily figure out how the graph would change and look. Have a look at this page it’s an interactive that explains enzymes quite well. http://www.wiley.com/college/boyer/0470003790/animations/enzyme_inhibition/enzyme_inhibition.htm
Disclaimer: The pictures and some text used are not of my own.
Enzymes Nomenclature or naming of enzymes:
There are 6 main classes of enzymes and each are assigned a number. From these classes are sub classes and sub sub-classes which are also assigned a number. When we lay out these numbers to which a particular enzyme belongs to we get what is known as the EC. Number or Enzyme Commission number and this was developed by the International Union of Biochemistry and Molecular Biology to properly classify enzymes in an orderly manner. Without further ado the following is a list of the major classes of enzymes and the number each class was assigned:
EC 1- Oxidoreductases
EC 2- Transferases
EC 3- Hydrolases
EC 4 –Lyases
EC 5 –Isomerases
EC 6 –Ligases
Now you might be thinking OMG this is soo hard to remember (i did to) but let’s just look at it its only 6 categories come on man you can learn that (what i told myself and i did). I am a strong advocate against acronyms because for; 1 they confuse you and you sometime don’t remember the original name of the think you are trying to remember, 2 if you can remember an acronym you can defiantly remember your work so stop that craziness and just learn you work… anyways.
Basically enzymes which catalyse oxidation reduction reactions, between substrates.
A– + B → A + B–
A is the reductant (electron donor) and B is the oxidant (electron acceptor)
Real life example: In glycolysis
Pi + glyceraldehyde-3-phosphate + NAD+ → NADH + H+ + 1,3-bisphosphoglycerate
NAD+ is the oxidant, and glyceraldehyde-3-phosphate is the reductant. This process is perfomed by the enzyme glyceraldehyde-3-phosphate dehygrogenase .
Catalyses the reaction which involve group transfer reactions. They allow the transfer of a from one molecule to another.
A–z + B → A + B–z
Real life example: In glycolysis
Glucose + ATPàglucose-6-phosphate + ADP
And this reaction is conducted by the enzyme hexokinase.
Brings about hydrolysis reactions with braking of bonds (using H2O)
A–B + H2O → A–OH + B–H
Example : nuclease is an enzyme capable of cleaving the phosphodiester bonds between the nucleotide subunits of nucleic acids.
Enzymes which catalyze the removal of group from substrates not using hydrolysis. Often forming a new double bond
Example: Decarboxylases that cleave carbon-carbon bonds
Enzymes act an inter-conversion of one optical of geometric or positional isomer to other.
Example: In glycolysis
Dihydroxyacetone phosphate àglyceraldehyde-3-phosphate
This reaction is catalysed by triose phosphate isomerise
Catalyse the reaction in which two compounds linked with breaking of phosphate bond in ATP of any similar compound providing energy.
Enzymes can also be divided by their specify:
Those that react with molecules with specific function groups example alcohol groups are said to exhibit group specify, some enzymes act on certain bonds/linkages such as amide linkages these are enzymes that are linkage specific while others react on optical and steric isomers which are called sterochemical specific enzymes.
Did you know that some fruits, vegetables and just general foods are great sources of these enzymes and we can observe the activity of these enzymes by simply using the tissue of such foods some common ones found in fruits and did you know that these enzymes can also help with our overall health? The following is a list of fruits and their enzymes put forward by www.everynutrient.com so feel free to have a look.
Pineapple (bromelain) – The bromelain in most digestive enzyme supplements is
extracted from pineapple stems, since they have the highest concentration of the
nutrient. The core and flesh of pineapple fruit contains good amounts of bromelain
as well. Bromelain is a group of powerful proteolytic digestive enzymes and provides
several other health benefits, most of which are still under investigation. Studies
have revealed that bromelain is also effective in fighting cancer growth. It blocks
growth of a broad range of tumor cells in several types of cancer including breast,
lung, colon, ovarian, and melanoma. Pineapple is also a great source of several
other nutrients including manganese, vitamin C, and potassium.
Green Papaya (papain) – Like the bromelain in pineapple, papain is a group of
proteolytic digestive enzymes. Papain, often extracted from papaya, is another
major ingredient in digestive enzyme supplements. Papain is also added to most
enzyme supplements that are formulated specifically for pain relief (arthritis, sports
injuries, etc.). Papain may also have anti-inflammatory properties. Papaya is an
excellent source of several other nutrients including potassium, calcium, vitamin C,
vitamin A, folate, beta-carotene, lutein, and zeaxanthin.
Mangoes (magneferin, katechol oxidase, and lactase) – In India, green mango
powder (amchur) is often used as a tenderizing agent for meats. Mango lassi is a
common drink in Indian restaurants and it’s made from a combination of mangoes,
yogurt, and spices. Not only are mangoes a rich source of digestive enzymes,
they’re also an excellent source of potassium, vitamin A, and beta-carotene.
Mangoes are also a good source of vitamin C, vitamin D, calcium, phosphorus,
magnesium, and fiber.
Kiwifruit (actinidin) – The actinidin enzyme in kiwifruit eases digestion due to it’s
proteolytic enzyme qualities. Actinidin is also found in pineapples, papayas, and
mangoes. Aside from kiwi being a great source of digestive enzymes, it’s also a
great source of several other nutrients including vitamin C (almost twice the amount
in an orange), magnesium, and potassium. Kiwi also acts as a blood thinner without
the adverse side effects of asprin.
Figs (ficin) – Used as a meat tenderizing agent, ficin is another protease (proteolytic)
enzyme that eases digestion. Ficin is found primarily in figs. Figs provide several
other health benefits as well. They’re an excellent source of fiber and a good source
of calcium, magnesium, and potassium.
Experiments and published papers coming soon along with structured multiple choice questions.
“So enzymes are biological catalysis that speeds up chemical reaction via providing an alternate pathway with lower activation energy.”JM
Remember from my protein post we can see that enzymes are proteins but are all enzymes proteins?
Nope…. some are RNA molecules called ribozymes and then we can also use some antibodies called abzymes (but also proteins).
Do we need a faster reaction why not just wait for it to happen naturally?
Well the answer to that is because you will die see some reaction within our bodies may take up to a trillion years if enzymes were not involved , according to Dr. Richard Wolfenden a distinguished professor of biochemistry, biophysics and chemistry at the University of North Carolina without a particular enzyme, a biological transformation he deemed “absolutely essential” in creating the building blocks of DNA and RNA would take 78 million years.
Can you believe that? Obviously we would be a cluster of carbon in the ground by that time however because of enzymes this reaction and many more can take place in a matter of milliseconds. Wolfenden also said “Without catalysts, there would be no life at all, from microbes to humans,” and this is certainly true.
We can show how enzymes work on a simple graph like the one below where we see how enzymes physically lowers the activation energy and as you should know the activation energy is the energy need to start the chemical reaction and a lower activation energy allows the reaction to take place faster.
So how exactly do enzymes work??
Enzymes attach to substrate via there active site and a molecule called an enzyme substrate complex is formed, this complex then quickly breaks down to form the product as well as the original enzyme. During this reaction bond or formed and broken and it is important to note that the enzyme substrate complex or intermediate has the highest energy since this is where the chemical reactions take place. We should also appreciate that there is no change to the enzyme, the enzyme remains the same after the reaction as it was before and when the product releases the enzyme moves on to another substrate molecule.
The active site is a cleft in the enzyme’s protein structure formed by the folding of amino acid residues, each active site is different this is because each enzyme contains a different protein structure and the properties of the active site that facilitate the combination of the substrate is due to the R groups of the proteins within the active site cleft, creating a complementary shape to the substrate that they bind to. Enzymes DO NOT make permanent bonds to substrate instead they bond via weak temporary bonds.
There are main theories about how enzymes and substrates react and bond:
1) Fischer’s lock and key hypothesis
This was developed by a scientist named Hermann Emil Fischer and he basically said that each enzyme it’s substrate fits perfectly together and complement each other forming the enzyme substrate complex and then the products which is a different shape from the substrate. The enzyme therefore acts as a lock and the substrate the key and only the key (substrate) for that lock (enzyme) would work or react to form a product. Once the products are formed they are released from the active site.
Although from this theory we can see that enzymes are highly specific and can only react with a certain substrate which must be perfectly complementary it is too strict as we know that this may not be the case in nature as most molecules are not perfect copies of each other thus another theory explains how enzymes react in this situation.
2) Koshland’s Induced fit hypothesis
This theory states that contrary to fischer’s theory all the enzyme’s active site and substrate are not perfectly complementary, they vary slightly however as the substrate approaches the enzyme changes the shape of its active site to fit the substrate and so the enzyme substrate complex is formed and the products are formed and released. This theory shows that enzymes are flexible and conformationaly dynamic molecules.
Michealis –Menten Curve
The reaction between enzymes and substrates can be expressed on a curve call a michealis –menten curve and this curve plots the velocity of the reaction against the substrate concentration (velocity is the amount of product formed per unit time or the amount of substrate being consumed). As the enzymes start to bind to the active sites the velocity increases and when all the active sites are occupied the graph remains constant, this point is called vmax and the enzymes are said to be saturated. This graph keeps all variables constant except the concentration of substrate. This graph forms a hyperbolic shape.
NOTE: This graph only shows the reaction for michealis menten enzymes of enzymes with one actives site present.
If an Allosteric enzyme were to be observed and plotted on the graph it would give a sigmoidal shape. Allosteric enzymes are those with more than one active site.
The black line shows the curve for an enzyme with one active site or a michealis –menten enzyme while the red show the curve for an allosteric enzyme.
This was the end of part one and honestly I never knew this much about enzymes who knew right… during cape (Alevels) we didn’t do much on enzymes unlike that of previous topics so this was all new to me but I coped well and I hope these notes help you . My next post will be a direct continuation from this and after that I will post 2 experiments I conducted recently …. Stay tuned hope you like 🙂