Several factors influence the rate at which an enzyme works. One of these is temperature, as the temperature increases so does the enzyme-catalysed reaction the reaction rate increases with temperature to a maximum level, then abruptly declines with further increase of temperature this is as the structure changes and the enzyme becomes denatured in fact most animal enzymes rapidly become denatured at temperatures above 40 C. The graph below shows this relationship:
The concentration of enzymes and substrates present can also influence the rate of reaction, the higher the concentration the faster the rate of reaction, the relationship is the same if you increase the amount of enzyme. Changes in pH also affect the rate of an enzyme reaction (how acidic or alkaline). Inhibitors like the other factors can influence the rate of reaction and are often used in industry and medicine. There are two common types of enzyme inhibition-competitive and non-competitive. Competitive inhibition is when an inhibitor similar to the substrate fits into the active site like a ‘key’ fits into a ‘lock’. The inhibitor competes for the active site and wins. Non-competitive inhibitors are considered to be substances which when added to the enzyme alter the enzyme in a way that it cannot accept the substrate. (Biology for You –Gareth Williams)
This essay will now look at particular enzymes in industry and medicine. An enzyme that is very important in the textile industry it is Amyloglucosidase, also known as glucoamylase. In the textile industry amylases are used to remove starch-based size for improved and uniform wet processing. The advantage of these enzymes is that they are specific for starch, removing it without damaging to the support fabric (e.g. cotton and its blends). Amyloglucosidase removes glucose residues in a stepwise manner from the non-reducing end of the starch polymer it hydrolyses the -1,4 and -1,6 bonds, although at slower rate with the -1,6 bond configuration. The amyloglucosidase enzymes can also catalyse the condensation of glucose residues producing mainly maltose and isomaltose. Glucoamylases are sensitive to temperature, being inactivated at temperatures above 60 ºC. However for this purpose this does not really affect its use. The optimum pH level of these enzymes is 4.0-4.5. This is important as if you know the optimum rate the enzyme functions quicker and then more of the product needed will be made quicker and therefore be able to be sold sooner. ()
Amyloglucosidase can be used with another enzyme to speed up the process of hydrolysis this enzyme is called Pullulanase. This is in the food industry however, here they are both co-immobilized using hydrophilic polyurethane foam. The combined amyloglucosidase and pullulanase activity of the immobilized enzyme was 32.2%+1.7% relative to the non-immobilized enzyme. The co-immobilized enzymes were capable of using a variety of glycogen and starch substrates. Co-immobilization of amyloglucosidase and pullulanase increased the glucose yield 1.6-fold over immobilized amyloglucosidase alone (). The co-immobilisation of these enzymes also gives a broader pH optimum from to pH 3 to 6.5 and gives increased stability at high temperatures as the it is not denatured until 80 C than the free enzymes but are more sensitive to inhibition by high KCl concentrations.
This is may be all well and good in the industry for making money, however some find it morally incorrect. This is as many people do not wish to have their food made through a process where the enzyme has been produced by a genetically engineered organism. Many feel strongly about this and have campaigned () to stop it as they feel enough is not known about the long-term effects of using them on humans and the ecosystem. Even though many campaign governments have released very little information on which enzymes are produced from genetically engineered organisms, they feel consumer confidence would lower and this cause problems for many large corporations and therefore cause problems for the economy. However due to this concern many food companies have launched an all organic range to appease people who feel this way and also so they do not miss out on this market which is increasingly growing as more research is done into this area.
Enzymes are increasingly used in the food industry one enzyme many are grateful for is Chymosin. This is actually one of the few enzymes that are known to have been made from genetically engineered organisms. Chymosin, known also as Rennin, is a proteolytic enzyme synthesized by chief cells in the stomach. Its role in digestion is to curdle or coagulate milk in the stomach, a process of considerable importance. If milk were not coagulated, it would rapidly flow through the stomach and miss the opportunity for initial digestion of its proteins. (http://arbl.cvmbs.colostate.edu/hbooks/pathphys/digestion/stomach/rennin.html)
However Chymosin is used in the food industry in cheese making. Chymosin efficiently converts liquid milk to a semisolid like cottage cheese, allowing it to be retained for longer periods. Its does the same in the body allowing it to be retained for longer periods in the stomach. In the body Chymosin secretion is maximal during the first few days after birth, and declines thereafter, replaced in effect by secretion of pepsin as the major gastric protease. Chymosin is also similar to pepsin in being most active in acidic environments, which makes sense considering its mission.
Chymosin’s structure consists of 323 amino acids in the polypeptide chain. The chain has three disulfide bridges. The molecule is kidney shaped and contains two domains. The folding pattern has pseudo 2-fold symmetry that is common for members of this acid proteinase family. (http://www.carb.nist.gov/gilliland_group/molecules/chymosin.html)
In order to understand how Chymosin is able to curdle milk one needs to come to an understanding of milk proteins. The majority of milk proteins is casein and there are four major types of casein molecules: alpha-s1, alpha-s2, beta and kappa. The alpha and beta caseins are hydrophobic proteins that are readily precipitated by calcium, the normal calcium concentration in milk is much more than that required to precipitate these proteins. However, kappa casein is a distinctly different molecule kappa casein it is not calcium precipitable. As the caseins are secreted, the alpha and beta caseins are kept from precipitating by their interactions with kappa casein. Simply what happens is that kappa casein normally keeps the majority of milk protein soluble and prevents it from suddenly coagulating, now enter the enzyme chymosin. Chymosin proteolytically cuts and inactivates kappa casein, converting it into para-kappa-casein and a smaller protein called macro peptide. Para-kappa-casein does not have the ability to stabilize the calcium-insoluble caseins precipitate, forming a curd. Chymosin at one time was the most widely used enzyme in cheese making and in days gone by, however today there are many other which at times are preferred to Chymosin such as some proteases which are able to coagulate milk by converting casein to para-casein. Even though discoveries like this have been made Chymosin is still an enzyme that is largely used. ()
Chymosin has however generated much controversy as it is an enzyme now known after much campaigning through the Freedom of Information act.() Many consumers may choose not to purchase cheese made using this enzyme as not enough is known about the long term effects of using an enzyme produced from a genetically engineered organism. Others argue that enzymes are usually found in minuscule quantities and by the time the final food product goes to market and so are not a danger. However many organisations such as the Organic Consumers Association believe that consumers should take care when buying there chosen food products as the long term effects are still a grey area. (Organic Consumers Association6101 Cliff Estate Rd., Little Marais, Minnesota 55614)
This essay will now look at enzymes used in medicine. Enzymes are used in all areas of medicine ranging from diagnosis to treatment. The enzyme first to be looked at is Streptokinase. In the past, management of thromboembolic vascular disease has relied on the use of anticoagulants, such as heparin and coumarin, to slow down the formation of fibrin clots. However, recognition that lysis of preformed fibrin could be accomplished by a process involving the conversion of inactive plasminogen to enzyme plasmin led to an alternative enzyme-based approach.
Then it was found that plasminogen activators, as opposed to preformed plasmin itself, can produce controlled enzymatic fibrinolysis. Two of these plasminogen activators are streptokinase and urokinase (enzymes). The mechanism of interaction between streptokinase and plasminogen is still argued over and remains undecided and a controversial issue. Native streptokinase inefficiently catalyses the conversion of human plasminogen to plasmin via the hydrolytic cleavage of a single L-arginyl-L-valyl bond. However, streptokinase is probably activated first by complexing with plasminogen in a one-to-one complex. This streptokinase-plasminogen complex in turn converts free plasminogen to plasmin with high efficiency. (http://www-biol.paisley.ac.uk/Courses/Enzymes/glossary/Therapy3.htm)
Artificially induced thrombi in the forearm veins of human volunteers were shown to be lysed by intravenously administered streptokinase. This enzyme has been shown to be of considerable use in the treatment of deep venous thrombosis and pulmonary embolism. It may also be of utility in the treatment of clotted hemodialysis shunts, priapism, and disseminated intravascular coagulation. However this enzyme is not as widely used a it could be this is due to the occurrence of rather frequent complications, allergic and anaphylactic reactions, Streptokinase has not gained the necessary approval and evaluation from the clinicists' side. With no definite reason, physicians try to avoid using Streptokinase, giving preference to urokinase and tissue plasminogen activator (t-PA), which are not endangered by allergic and anaphylactic complications, but are expensive and not available to broader public. However those who are not as well off have to be treated using Streptokinase and they can be treated not worse than those who are well off, the only difference being the length of treatment which on average is by 30-35 days longer, most often at expense of Streptokinase desensibilization course. ()
Many other enzymes are used in medicine and for many other purposes. One purpose is when diabetics need to measure their glucose levels, this is an analytical test. This is always measured by an enzyme based test utilising glucose oxidase. Diabetics use strips of paper impregnated with glucose oxidase to monitor their blood sugar. When a drop of blood is added to the strip, the glucose oxidase metabolises the glucose and a series of reactions produce a measurable colour change that is proportional to the amount of glucose. ()
Enzymes are actually used as medicines, usually to replace enzyme deficiencies in patients. An example is the use of blood clotting factors to treat haemophiliacs, or the opposite where proteases are used to degrade fibrin; use of proteases can prevent the formation of dangerous blood clots. The protease (peptidase) used in therapy of thromboembolic diseases (myocardial infarction, embolisms and deep venous thrombosis) is called tissue plasminogen activator. Nuclease is also considered as a possible therapy for cystic fibrosis, but it is not clear how commercialised and therapeutically successful this has been. (Medical Journal Article - Dr. Pavels Ivdra - 1996)
Proteases are also used in wound therapy. In this case they are called debriding agents and are used to clean a wound and therefore accelerate the healing process.
Some proteases are also used as anti-inflammatory reagents. An enzyme called super oxide dismutase is also available as an anti-inflammatory agent, but how successful it has been as a commercial product, is not clear but medically the product does the job required. (www.enzymes.co.uk)
Enzymes can also be used in a healing way by aiding bodily functions. Enzymes are used to aid digestion, both in humans and animals. In humans, enzymes are used to supplement the natural amylase, lipase and protease produced normally by the pancreas. Many people also have a problem called lactose intolerance. This means as these people get older they lose the enzyme lactase (converts lactose into glucose and galactose). This means they cannot ingest milk or dairy products. Lactase supplements help to avoid stomach upsets for these people. (Biology for You – Gareth Williams)
In conclusion it is simple to see that life today as we know would be very different, in fact it would be impossible as enzymes are used everyday by our bodies to perform what some may see as everyday functions. However in regards to industry and medicine in particular enzymes play a huge role. They are used in many of the treatments administered to patients and much more. They also play a major role in industry, becoming over the years a billion dollar market. Quite simply although they go unnoticed by many they are owned a great amount of gratitude and as a great immunologist Dr. Pavels Ivdra once said ‘Enzymes the unsung heroes’ (Medical Journal 1996).
Bibliography:
Collins Advanced Sciences Biology - Various authors
Biology for You –Gareth Williams
Organic Consumers Association6101 Cliff Estate Rd., Little Marais, Minnesota 55614
Medical Journal Article - Dr. Pavels Ivdra – 1996