Bioenergetics  and Enzymes
Bioenergetics
<ul><li>Act as  catalysts  for metabolic reactions </li></ul><ul><li>Are  globular proteins  with specific 3D formations <...
Enzymes <ul><li>Enzymes are the  catalysts  of metabolic rxns. </li></ul><ul><ul><li>Catalyst  – affects a chemical reacti...
Mechanisms of Enzyme Action
Lock & Key Model <ul><li>Enzyme – Substrate Specificity! </li></ul>Substrate Enzyme Active Site
<ul><li>Usually large globular proteins (a) </li></ul><ul><li>b)  Active Site  where the substrate combines to the enzyme ...
The  induced fit  between an enzyme and its substrate
<ul><li>Active site is not a rigid pocket for the substrate to fit in. </li></ul><ul><li>Substrate “induces” the enzyme to...
The catalytic cycle of an enzyme
Inhibitors <ul><li>General function: </li></ul><ul><ul><li>Reduce activity of enzyme </li></ul></ul><ul><ul><li>Stop enzym...
Metabolic Pathways <ul><li>Metabolic Pathways </li></ul>
 
 
Competitive Inhibitor <ul><li>Very similar to the substrate </li></ul><ul><li>Compete with the substrate on the active sit...
 
 
Competitive Inhibitor - Example <ul><li>Krebs Cycle </li></ul><ul><li>Malonate – inhibitor of cell respiration </li></ul><...
Krebs's Cycle – A METABOLIC PATHWAY
Non - Competitive <ul><li>Molecules NOT similar to the substrate </li></ul><ul><ul><li>NO competition for the active site ...
 
TWO TYPES
Allosteric
Allsoteric <ul><li>FEEDBACK INHIBITION  </li></ul>
Allosteric Inhibitors <ul><li>Non competitive (end – product inhibition) </li></ul><ul><li>Bind to a site in the enzyme:  ...
 
Allosteric Inhibitors <ul><li>“feedback inhibition” </li></ul><ul><li>“ End product inhibition”: the last product of a rea...
Example  <ul><li>ATP – Feedback inhibition  (during glycolysis) </li></ul><ul><li>ATP accumulates </li></ul><ul><li>Acts a...
Irreversible inhibitors <ul><li>Generally are poisonous substances that enter from the outside of the body. </li></ul><ul>...
Inhibition Animations <ul><li>ACE Inhibitors </li></ul><ul><li>The Science of Enzyme Inhibition </li></ul>
Enzyme Action and Energy
How Do Enzymes Work? <ul><li>For a reaction to start it needs energy: </li></ul><ul><li>ACTIVATION ENERGY </li></ul><ul><l...
Energy Progress of Rxn. Reactants Products
Energy Progress of Rxn. Reactants Products Activation Energy - Minimum energy to make the reaction happen
Energy Progress of Rxn. Reactants Products Enzyme Substrate Complex or Transition State
Energy Progress of Rxn. Reactants Products Overall energy change
Energy Progress of Rxn. Reactants Products Exergonic
Enzymes and Reactions <ul><li>Reactions are not impossible without enzymes. </li></ul><ul><ul><li>With enzymes the RATE of...
Energy in a Reaction <ul><li>Reactants must absorb energy from their surroundings for their bonds to break. </li></ul><ul>...
Endergonic vs. Exergonic
Endergonic Reaction <ul><li>Endergonic Reaction  (Endothermic)  –  </li></ul><ul><ul><li>Absorb energy into the reaction <...
Endergonic Rxn.
Exergonic Reaction <ul><li>Give off Energy </li></ul><ul><ul><li>Represented by a negative change in energy </li></ul></ul...
Energy profile of an exergonic reaction
Figure 6.13  Enzymes lower the barrier of activation energy
Catalyzed Reactions <ul><li>The Small Intestine Reactions: </li></ul><ul><li>Starch  +  Amylase   = many maltose units </l...
A Cell Reaction <ul><li>Cytotoxic Reaction: </li></ul><ul><li>2H 2 O 2   + catalase     O 2  + 2H 2 O </li></ul><ul><li>G...
Figure 6.15  The catalytic cycle of an enzyme
Characteristics of Enzyme Activity <ul><li>1.  Enzymes work best at certain temperatures and enable cell reactions to proc...
Characteristics of Enzyme Activity <ul><li>THE MAD GOOD ANNY’ </li></ul>
Figure 6.16  Environmental factors affecting enzyme activity
Temperature <ul><li>As you  increase   temperature ,  enzyme action   increases  as well until an optimum temperature for ...
Enzymes are temperature dependent…... (in humans) <ul><li>Most work at body temperature:37 o C to maintain homeostasis </l...
Effect of  Substrate Concentration <ul><li>As substrate concentration increases, the rate of enzyme catalyzed reactions wi...
pH <ul><li>Affects enzyme action. </li></ul><ul><li>Certain enzymes work better in acidic environments while others in bas...
Effect of  pH  on Enzyme Activity <ul><li>Enzymes have an optimum pH at which they function best. </li></ul><ul><li>At oth...
Enzymes are pH specific. <ul><li>Different enzymes </li></ul><ul><li>Different body areas </li></ul><ul><li>Different opti...
 
Co-existing with Enzymes: <ul><li>Cofactors and Coenzymes </li></ul>
Cofactors <ul><li>Needs to be present in addition to an enzyme for a reaction to catalyze. </li></ul><ul><li>Like enzymes,...
COENZYMES <ul><li>A type of cofactor ---- ORGANIC in nature </li></ul><ul><ul><ul><li>Separate from the protein in the enz...
COENZYMES
Application of Enzymes in Biotechnology
<ul><li>Pectinase – ******IB Req. </li></ul><ul><li>Pectin – polysaccharide found in fruit skins (cell wall) </li></ul><ul...
Lactose intolerance <ul><li>Unable to produce </li></ul><ul><li>lactase in sufficient </li></ul><ul><li>quantities. </li><...
<ul><li>Biological  Detergents </li></ul><ul><li>Proteases – protein (most dirt) </li></ul><ul><li>Source: Bacteria – B ac...
Making Cheese <ul><li>Rennin – an enzyme that turns milk to cheese. </li></ul><ul><li>Coagulates into curd at body tempera...
Milk <ul><li>Adding LACTASE to milk. </li></ul><ul><li>Breaks down LACTOSE into easier digestible sugars </li></ul><ul><li...
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Bioenergentics and Enzymes III

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  • Bioenergetics, loosely defined, is the study of energy investment and flow through living systems. This broad definition includes the study of thousands of different processes ranging from cellular respiration and the production of ATP , to the study of evolutionary costs accompanying the development of a particular trait, such as the immune system . One question this area of science seeks to answer is whether protective benefit of a particular trait is worth the energy investment it requires .
  • enzyme : a globular protein functioning as a biological catalyst, speeding up reaction rates by lowering activation energy active site : the site on the surface of an enzyme to which substrate(s) bind Substrate – the molecule on which the enzyme acts
  • enzyme : a globular protein functioning as a biological catalyst, speeding up reaction rates by lowering activation energy active site : the site on the surface of an enzyme to which substrate(s) bind
  • Explain enzyme-substrate specificity each globular enzyme includes an active site with a specific, three-dimensional shape which is complementary to the shape of the substrate the globular enzyme active site also includes a specific set of charges which are complementary to the charges of the substrate thus, through complementarity of shape and charge, the substrate is attracted to, and fits precisely into, the active site the precise interactions between enzyme active site and substrate are essential for the catalytic properties of enzymes to funciton; the complementarity is often referred to as analogous to the fit between a lock and a key enzymes vary in specificity from being exclusive to a single substrate to being generalized to accept any molecule of a certain type
  • each globular enzyme includes an active site with a specific, three-dimensional shape which is complementary to the shape of the substrate the globular enzyme active site also includes a specific set of charges which are complementary to the charges of the substrate thus, through complementarity of shape and charge, the substrate is attracted to, and fits precisely into, the active site the precise interactions between enzyme active site and substrate are essential for the catalytic properties of enzymes to funciton; the complementarity is often referred to as analogous to the fit between a lock and a key enzymes vary in specificity from being exclusive to a single substrate to being generalized to accept any molecule of a certain type
  • each globular enzyme includes an active site with a specific, three-dimensional shape which is complementary to the shape of the substrate the globular enzyme active site also includes a specific set of charges which are complementary to the charges of the substrate thus, through complementarity of shape and charge, the substrate is attracted to, and fits precisely into, the active site the precise interactions between enzyme active site and substrate are essential for the catalytic properties of enzymes to funciton; the complementarity is often referred to as analogous to the fit between a lock and a key enzymes vary in specificity from being exclusive to a single substrate to being generalized to accept any molecule of a certain type
  • For many enzymes, the lock-and-key model does not fully explain the binding of the substrate to the active site As the substrate approaches the active site and binds to it, the shape of the active site changes and only then does the active site conform, and become complementary to fit the shape of the substrate The substrate induces the active site to change, weakening bonds in the substrate during the process, and thus reducing activation energy The induced fit model helps explain the broad specificity of some enzymes
  • For many enzymes, the lock-and-key model does not fully explain the binding of the substrate to the active site As the substrate approaches the active site and binds to it, the shape of the active site changes and only then does the active site conform, and become complementary to fit the shape of the substrate The substrate induces the active site to change, weakening bonds in the substrate during the process, and thus reducing activation energy The induced fit model helps explain the broad specificity of some enzymes
  • State that metabolic pathways consist of chains and cycles of enzyme catalyzed reactions Describe the induced fit model For many enzymes, the lock-and-key model does not fully explain the binding of the substrate to the active site As the substrate approaches the active site and binds to it, the shape of the active site changes and only then does the active site conform, and become complementary to fit the shape of the substrate The substrate induces the active site to change, weakening bonds in the substrate during the process, and thus reducing activation energy The induced fit model helps explain the broad specificity of some enzymes
  • competitive inhibition substrate and inhibitor are chemically very similar inhibitor binds to the active site of the enzyme while the inhibitor occupies the active site, it prevents the substrate from binding, and so the activity of the enzyme is prevented until the inhibitor dissociates example: folic acid synthetase folic acid synthetase is an enzyme in bacteria which normally produces folic acid, an essential vitamin, from PABA and other substrates a group of antibiotics, known as sulfanilamides, binds to and occupies the active site of folic acid synthetase, thus blocking the access of the similarly shaped substrate, PABA without folic acid, the bacteria die, and the infection is overcome non-competitive inhibition substrate and inhibitor are not similar inhibitor binds to the enzyme at a site different than the active site the inhibitor changes the conformation (3-D, tertiary structure) of the enzyme, causing enough alteration to slow enzyme activity; while substrate binds to the active site, it is not converted to a product example: silver, Ag+ silver forms bonds with the -SH groups of cysteine, the amino acid which normally forms covalent disulfide bridges the disruption of disulfide bridges alters the tertiary structure of the enzyme, affecting its active site thus, silver (and other heavy metals) act as metabolic poisons by disrupting the activity of many enzymes
  • competitive inhibition substrate and inhibitor are chemically very similar inhibitor binds to the active site of the enzyme while the inhibitor occupies the active site, it prevents the substrate from binding, and so the activity of the enzyme is prevented until the inhibitor dissociates example: folic acid synthetase folic acid synthetase is an enzyme in bacteria which normally produces folic acid, an essential vitamin, from PABA and other substrates a group of antibiotics, known as sulfanilamides, binds to and occupies the active site of folic acid synthetase, thus blocking the access of the similarly shaped substrate, PABA without folic acid, the bacteria die, and the infection is overcome non-competitive inhibition substrate and inhibitor are not similar inhibitor binds to the enzyme at a site different than the active site the inhibitor changes the conformation (3-D, tertiary structure) of the enzyme, causing enough alteration to slow enzyme activity; while substrate binds to the active site, it is not converted to a product example: silver, Ag+ silver forms bonds with the -SH groups of cysteine, the amino acid which normally forms covalent disulfide bridges the disruption of disulfide bridges alters the tertiary structure of the enzyme, affecting its active site thus, silver (and other heavy metals) act as metabolic poisons by disrupting the activity of many enzymes
  • competitive inhibition substrate and inhibitor are chemically very similar inhibitor binds to the active site of the enzyme while the inhibitor occupies the active site, it prevents the substrate from binding, and so the activity of the enzyme is prevented until the inhibitor dissociates example: folic acid synthetase folic acid synthetase is an enzyme in bacteria which normally produces folic acid, an essential vitamin, from PABA and other substrates a group of antibiotics, known as sulfanilamides, binds to and occupies the active site of folic acid synthetase, thus blocking the access of the similarly shaped substrate, PABA without folic acid, the bacteria die, and the infection is overcome
  • folic acid synthetase is an enzyme in bacteria which normally produces folic acid, an essential vitamin, from PABA and other substrates a group of antibiotics, known as sulfanilamides, binds to and occupies the active site of folic acid synthetase, thus blocking the access of the similarly shaped substrate, PABA without folic acid, the bacteria die, and the infection is overcome
  • non-competitive inhibition substrae and inhibitor are not similar inhibitor binds to the enzyme at a site different than the active site the inhibitor changes the conformation (3-D, tertiary structure) of the enzyme, causing enough alteration to slow enzyme activity; while substrate binds to the active site, it is not converted to a product example: silver, Ag+ silver forms bonds with the -SH groups of cysteine, the amino acid which normally forms covalent disulfide bridges the disruption of disulfide bridges alters the tertiary structure of the enzyme, affecting its active site thus, silver (and other heavy metals) act as metabolic poisons by disrupting the activity of many enzymes
  • example: silver, Ag+ silver forms bonds with the -SH groups of cysteine, the amino acid which normally forms covalent disulfide bridges the disruption of disulfide bridges alters the tertiary structure of the enzyme, affecting its active site thus, silver (and other heavy metals) act as metabolic poisons by disrupting the activity of many enzymes Hg 2+ , Ag + , Cu 2+ , CN- bind to to SH groups and break di sulfide bridge (cytochrome oxidase) NERVE GAS – SARIN – inhibit acetyl cholinesterase
  • Another form of inhibition involves an inhibitor that binds to an allosteric site of an enzyme.  An allosteric site is a different location than the active site. The binding of an inhibitor to the allosteric site alters the shape of the enzyme, resulting in a distorted active site that does not function properly. The binding of an inhibitor to an allosteric site is usually temporary.   Poisons are inhibitors that bind irreversibly. For example, penicillin inhibits an enzyme needed by bacteria to build the cell wall.
  • When ATP accumulates, it acts an allosteric inhibitor of the enzyme phosphofructokinase. THIS LOWERS THE RATE OF REACTION AND LESS ATP IS PRODUCED. ATP PRODUCTION IS CONTROLLED BY ATP Feedback inhibition – atp production inhibits the production of the enzyme which then inhibits the production of the ATP to prevent accumulation Feedback Inhibition Negative feedback inhibition is like a thermostat. When it is cold, the thermostat turns on a heater which produces heat. Heat causes the thermostat to turn off the heater. Heat has a negative effect on the thermostat; it feeds back to an earlier stage in the control sequence as diagrammed below.  Many enzymatic pathways are regulated by feedback inhibition. As an enzyme&apos;s product accumulates, it turns off the enzyme just as heat causes a thermostat to turn off the production of heat.  The end product of the pathway binds to an allosteric site on the first enzyme in the pathway and shuts down the entire sequence.    
  • competitive inhibition substrate and inhibitor are chemically very similar inhibitor binds to the active site of the enzyme while the inhibitor occupies the active site, it prevents the substrate from binding, and so the activity of the enzyme is prevented until the inhibitor dissociates example: folic acid synthetase folic acid synthetase is an enzyme in bacteria which normally produces folic acid, an essential vitamin, from PABA and other substrates a group of antibiotics, known as sulfanilamides, binds to and occupies the active site of folic acid synthetase, thus blocking the access of the similarly shaped substrate, PABA without folic acid, the bacteria die, and the infection is overcome non-competitive inhibition substrate and inhibitor are not similar inhibitor binds to the enzyme at a site different than the active site the inhibitor changes the conformation (3-D, tertiary structure) of the enzyme, causing enough alteration to slow enzyme activity; while substrate binds to the active site, it is not converted to a product example: silver, Ag+ silver forms bonds with the -SH groups of cysteine, the amino acid which normally forms covalent disulfide bridges the disruption of disulfide bridges alters the tertiary structure of the enzyme, affecting its active site thus, silver (and other heavy metals) act as metabolic poisons by disrupting the activity of many enzymes
  • Cytochrome oxidase - aerobic respiration in the mitochondria =…suffocates The disruption is caused by blocking acetylcholinesterase , an enzyme that normally relaxes the activity of acetylcholine , a neurotransmitter
  • chemical reaction reactants converted into products activation energy must be exceeded for any reaction to occur allows breaking of bonds in exergonic reactions allows formation of new bonds in endergonic reactions enzymes reduce activation energy active site interacts with substrate, altering stability of substrate bonds, allowing substrate molecules to form a transition state which is different than would be formed without the enzyme transition state of enzyme-catalyzed reactions has lower energy than non-enzyme-catalyzed reactions, thus lowering activation energy the net energy released in exergonic reactions, or taken in by endergonic reactions, is not changed by the enzyme, which only reduces activation energy by lowering the energy requirements of the transition state of reactants
  • chemical reaction reactants converted into products activation energy must be exceeded for any reaction to occur allows breaking of bonds in exergonic reactions allows formation of new bonds in endergonic reactions enzymes reduce activation energy active site interacts with substrate, altering stability of substrate bonds, allowing substrate molecules to form a transition state which is different than would be formed without the enzyme transition state of enzyme-catalyzed reactions has lower energy than non-enzyme-catalyzed reactions, thus lowering activation energy the net energy released in exergonic reactions, or taken in by endergonic reactions, is not changed by the enzyme, which only reduces activation energy by lowering the energy requirements of the transition state of reactants
  • chemical reaction reactants converted into products activation energy must be exceeded for any reaction to occur allows breaking of bonds in exergonic reactions allows formation of new bonds in endergonic reactions enzymes reduce activation energy active site interacts with substrate, altering stability of substrate bonds, allowing substrate molecules to form a transition state which is different than would be formed without the enzyme transition state of enzyme-catalyzed reactions has lower energy than non-enzyme-catalyzed reactions, thus lowering activation energy the net energy released in exergonic reactions, or taken in by endergonic reactions, is not changed by the enzyme, which only reduces activation energy by lowering the energy requirements of the transition state of reactants
  • chemical reaction reactants converted into products activation energy must be exceeded for any reaction to occur allows breaking of bonds in exergonic reactions allows formation of new bonds in endergonic reactions enzymes reduce activation energy active site interacts with substrate, altering stability of substrate bonds, allowing substrate molecules to form a transition state which is different than would be formed without the enzyme transition state of enzyme-catalyzed reactions has lower energy than non-enzyme-catalyzed reactions, thus lowering activation energy the net energy released in exergonic reactions, or taken in by endergonic reactions, is not changed by the enzyme, which only reduces activation energy by lowering the energy requirements of the transition state of reactants
  • chemical reaction reactants converted into products activation energy must be exceeded for any reaction to occur allows breaking of bonds in exergonic reactions allows formation of new bonds in endergonic reactions enzymes reduce activation energy active site interacts with substrate, altering stability of substrate bonds, allowing substrate molecules to form a transition state which is different than would be formed without the enzyme transition state of enzyme-catalyzed reactions has lower energy than non-enzyme-catalyzed reactions, thus lowering activation energy the net energy released in exergonic reactions, or taken in by endergonic reactions, is not changed by the enzyme, which only reduces activation energy by lowering the energy requirements of the transition state of reactants
  •   Laws of thermodynamics    1.         Energy can be transferred and transformed but neither created nor destroyed     2.         Every energy transfer or transformation increases the entropy of the universe. Energy lost to entropy is usually in the form of heat ( i.e. , the random movement of particles). When we eat food we convert ordered forms of energy (carbohydrates, proteins, and lipids) into disordered forms (heat).   B.  Free energy (G) - the portion of a system’s energy that is available to do work                          1.   Two types of processes exist: spontaneous and nonspontaneous. Spontaneous reactions are those that occur without outside help and can be used to perform work. When spontaneous reactions occur, the system becomes more stable, and the entropy of the universe increases.                           2.     Basic relationship is  order =  energy =  instability (or  stability) =  entropy                         3.  Systems with high energy are unstable and prefer to lose that energy. The prefer to go from ordered to less ordered. To be spontaneous, free energy must be lost. The system must lose energy (decrease in H), gain entropy (increase in S), or both. When these two are added, ΔG must be negative.                           4.  The greater the decrease in G, the more work can be done.        
  • chemical reaction reactants converted into products activation energy must be exceeded for any reaction to occur allows breaking of bonds in exergonic reactions allows formation of new bonds in endergonic reactions enzymes reduce activation energy active site interacts with substrate, altering stability of substrate bonds, allowing substrate molecules to form a transition state which is different than would be formed without the enzyme transition state of enzyme-catalyzed reactions has lower energy than non-enzyme-catalyzed reactions, thus lowering activation energy the net energy released in exergonic reactions, or taken in by endergonic reactions, is not changed by the enzyme, which only reduces activation energy by lowering the energy requirements of the transition state of reactants
  • Hydrogen peroxide is a liquid made up of two atoms of hydrogen and two oxygen atoms (H2O2).  As a molecule, it is similar in structure to water (H2O), but less stable.  It readily breaks down into water and oxygen when placed in contact with something it can react with.  For example, if you pour hydrogen peroxide on a wound, you will see a fizzle similar to bubbles that appear when soda pop is poured into a glass.  Those bubbles on the wound are oxygen.  Hydrogen peroxide is not only found at the drug store...it is also produced in the human body by cells of the immune system, for example.  These cells (eg. neutrophils) make H2O2 to combat infection during the inflammatory process.  Hydrogen peroxide kills cells by disrupting/destroying their cell membranes.  This is a non-specific process.  In other words, H2O2 can, and does, cause harm to the human body.  The biochemistry of hydrogen peroxide is complex and widely researched.  It is an essential molecule for our survival.  However, our bodies are pretty smart; they use this reactive molecule under controlled conditions to prevent damaging normal structures.  As a liquid, hydrogen peroxide is used in conventional western medicine and alternative medicine to treat patients.  In conventional medicine, it is mixed with sterile water and used topically (on the skin) to wash infected or dirty wounds.  It is also an ingredient of dental whitening kits and strips.  Carbonic Acid is then broken into the carbonate ion and a hydrogen ion. Carbonic anhydrase works at great speeds…gas exchange
  • reasons why enzymes are altered by environmental conditions each enzyme has a highly specifically shaped active site which is complementary to the shape of its substrate; catalysis depends on this complementarity the enzyme active site is a product of its tertiary, or three-dimensional structure, which is in turn produced by a variety of bonds: covalent, ionic , and hydrogen bonds , as well as hydrophobic interactions each enzyme active site best fits its substrate at a set of optimum conditions deviation from optimum conditions alter the bonds which produce the tertiary structure of the enzyme, thus altering the shape of the active site and its complementary fit to its substrate pH both acids and alkalis denature enzymes stomach pepsin is optimized at pH=2 pancreatic lipase is optimized at pH=8 temperature at lower temperatures, all chemical reactions proceed more slowly, with a general rule of doubling reaction rates with each 10 degrees Celcius increase at higher temperatures, the excessive energy breaks bonds that would otherwise create the shape of the active site; this denaturing the enzyme substrate concentration at low to medium substrate concentrations, enzyme activity is directly proportional to substrate concentration; this is because random collisions between substrate and active site happen more frequently with higher substrate concentrations at high substrate concentrations, all the active sites of the enzymes are fully occupied, so raising the substrate concentration has no effect
  • reasons why enzymes are altered by environmental conditions each enzyme has a highly specifically shaped active site which is complementary to the shape of its substrate; catalysis depends on this complementarity the enzyme active site is a product of its tertiary, or three-dimensional structure, which is in turn produced by a variety of bonds: covalent, ionic , and hydrogen bonds , as well as hydrophobic interactions each enzyme active site best fits its substrate at a set of optimum conditions deviation from optimum conditions alter the bonds which produce the tertiary structure of the enzyme, thus altering the shape of the active site and its complementary fit to its substrate pH both acids and alkalis denature enzymes stomach pepsin is optimized at pH=2 pancreatic lipase is optimized at pH=8 temperature at lower temperatures, all chemical reactions proceed more slowly, with a general rule of doubling reaction rates with each 10 degrees Celcius increase at higher temperatures, the excessive energy breaks bonds that would otherwise create the shape of the active site; this denaturing the enzyme substrate concentration at low to medium substrate concentrations, enzyme activity is directly proportional to substrate concentration; this is because random collisions between substrate and active site happen more frequently with higher substrate concentrations at high substrate concentrations, all the active sites of the enzymes are fully occupied, so raising the substrate concentration has no effect
  • reasons why enzymes are altered by environmental conditions each enzyme has a highly specifically shaped active site which is complementary to the shape of its substrate; catalysis depends on this complementarity the enzyme active site is a product of its tertiary, or three-dimensional structure, which is in turn produced by a variety of bonds: covalent, ionic , and hydrogen bonds , as well as hydrophobic interactions each enzyme active site best fits its substrate at a set of optimum conditions deviation from optimum conditions alter the bonds which produce the tertiary structure of the enzyme, thus altering the shape of the active site and its complementary fit to its substrate pH both acids and alkalis denature enzymes stomach pepsin is optimized at pH=2 pancreatic lipase is optimized at pH=8 temperature at lower temperatures, all chemical reactions proceed more slowly, with a general rule of doubling reaction rates with each 10 degrees Celcius increase at higher temperatures, the excessive energy breaks bonds that would otherwise create the shape of the active site; this denaturing the enzyme substrate concentration at low to medium substrate concentrations, enzyme activity is directly proportional to substrate concentration; this is because random collisions between substrate and active site happen more frequently with higher substrate concentrations at high substrate concentrations, all the active sites of the enzymes are fully occupied, so raising the substrate concentration has no effect
  • reasons why enzymes are altered by environmental conditions each enzyme has a highly specifically shaped active site which is complementary to the shape of its substrate; catalysis depends on this complementarity the enzyme active site is a product of its tertiary, or three-dimensional structure, which is in turn produced by a variety of bonds: covalent, ionic , and hydrogen bonds , as well as hydrophobic interactions each enzyme active site best fits its substrate at a set of optimum conditions deviation from optimum conditions alter the bonds which produce the tertiary structure of the enzyme, thus altering the shape of the active site and its complementary fit to its substrate substrate concentration at low to medium substrate concentrations, enzyme activity is directly proportional to substrate concentration; this is because random collisions between substrate and active site happen more frequently with higher substrate concentrations at high substrate concentrations, all the active sites of the enzymes are fully occupied, so raising the substrate concentration has no effect
  • reasons why enzymes are altered by environmental conditions each enzyme has a highly specifically shaped active site which is complementary to the shape of its substrate; catalysis depends on this complementarity the enzyme active site is a product of its tertiary, or three-dimensional structure, which is in turn produced by a variety of bonds: covalent, ionic , and hydrogen bonds , as well as hydrophobic interactions each enzyme active site best fits its substrate at a set of optimum conditions deviation from optimum conditions alter the bonds which produce the tertiary structure of the enzyme, thus altering the shape of the active site and its complementary fit to its substrate pH both acids and alkalis denature enzymes stomach pepsin is optimized at pH=2 pancreatic lipase is optimized at pH=8
  • A cofactor is any substance that needs to be present in addition to an enzyme to catalyze a certain reaction. (However, more or less ubiquitous substances such as water do not qualify.) Some cofactors are inorganic , such as the metal atoms zinc , magnesium, iron , and copper in certain forms. Others, such as most vitamins , are organic, and are known as coenzymes . Some cofactors undergo chemical changes during the course of a reaction (i.e. being reduced or oxidized ). Nonetheless, as a catalyst , cofactors will be returned to their original state when the reaction in which they are needed has finished -- they are not consumed in the reaction or permanently converted to something else (that would be a substrate of the reaction). Cofactors vary in location and tightness of binding. When bound tightly to the enzyme, they are called prosthetic groups . Loosely bound cofactors typically bind in a similar fashion to enzyme substrates . When a cofactor is an organic substance that directly participates as a substrate in the reaction, it is called a coenzyme . Vitamins can serve as precursors to coenzymes (e.g. vitamins B1 , B2 , B6 , B12 , niacin , folic acid ) or as cofactors themselves (e.g. vitamin C ). Cofactors are inorganic ions and organic, non-protein molecules that help some enzymes function as catalysts. When inorganic, they are usually either copper, zinc or iron. Found on the active sites of enzymes, they attract electrons from bonds in a substrate to cause them to break.
  • A cofactor is any substance that needs to be present in addition to an enzyme to catalyze a certain reaction. (However, more or less ubiquitous substances such as water do not qualify.) Some cofactors are inorganic , such as the metal atoms zinc , magnesium, iron , and copper in certain forms. Others, such as most vitamins , are organic, and are known as coenzymes . Some cofactors undergo chemical changes during the course of a reaction (i.e. being reduced or oxidized ). Nonetheless, as a catalyst , cofactors will be returned to their original state when the reaction in which they are needed has finished -- they are not consumed in the reaction or permanently converted to something else (that would be a substrate of the reaction). Cofactors vary in location and tightness of binding. When bound tightly to the enzyme, they are called prosthetic groups . Loosely bound cofactors typically bind in a similar fashion to enzyme substrates . When a cofactor is an organic substance that directly participates as a substrate in the reaction, it is called a coenzyme . Vitamins can serve as precursors to coenzymes (e.g. vitamins B1 , B2 , B6 , B12 , niacin , folic acid ) or as cofactors themselves (e.g. vitamin C ). Cofactors are inorganic ions and organic, non-protein molecules that help some enzymes function as catalysts. When inorganic, they are usually either copper, zinc or iron. Found on the active sites of enzymes, they attract electrons from bonds in a substrate to cause them to break.
  • pectinase in fruit juice production pectin is a complex polysaccharide, found in the cell walls of plants; pectinase is an enzyme that breaks down pectin by hydrolysis reactions pectinase is obtained by artificially culturing a fungus, Aspergillus niger ; the fungus grows naturally on fruits where it uses pectinase to soften the cell walls of the fruit so that it can grow through it fruit juices are produced by crushing ripe fruits to separate liquid juice from solid pulp; when ripe fruits are crushed, pectin forms links between the cell wall and the cytoplasm of the fruit cells, making the juice viscous and more difficult to separate from the pulp; pectinase is added during crushing of fruit of fruit to break down the pectin pectinase makes juice more fluid and easy to separate from the pulp; it therefore increases the volume of juice that is obtained; it also makes the juice less cloudy by helping solids suspended in the juice to settle and be separated from the liquid
  • protease in biological washing powder protease enzymes break down proteins into soluble peptides and amino acids; laundry washing powders that contain protease are called biological washing powders protease is obtained by culturing a bacterium, Bacillus licheniformis , that is adapted to grow in alkaline conditions; this bacterium feeds on proteins in its habitat by secreting protease; the protease has a high pH optimum of between 9 and 10 detergents in laundry washing powders remove fats and oils during the washing of clothes, but much of the dirt on clothing is made of protein, not lipids; if protease is added to the washing powder, this protein is digested during the wash; the high pH optimum of protease allows it to remain active, despite the high pH caused by alkalis in the washing powder if protease is not used, protein stains on clothes can only be removed by using a very high temperature wash; protease allows much lower temperatures to be used, with lower energy use and less risk of shrinkage of garments or loss of colored dyes
  • Bioenergentics and Enzymes III

    1. 1. Bioenergetics and Enzymes
    2. 2. Bioenergetics
    3. 3. <ul><li>Act as catalysts for metabolic reactions </li></ul><ul><li>Are globular proteins with specific 3D formations </li></ul><ul><li>Have a high specificity for substrates </li></ul><ul><li>Substrate – the molecule on which the enzyme acts. </li></ul><ul><li>Possess an “ active site ” on the surface of the enzyme in which substrates readily interact </li></ul>Enzymes How Enzymes Work How Enzymes Work
    4. 4. Enzymes <ul><li>Enzymes are the catalysts of metabolic rxns. </li></ul><ul><ul><li>Catalyst – affects a chemical reaction without itself being changed (re-used). </li></ul></ul><ul><li>Enzymes are large PROTEINS! </li></ul><ul><li>Most enzymes end in “ ase” . </li></ul><ul><li>Enzymes regulate the rate at which reactions occur. </li></ul>
    5. 5. Mechanisms of Enzyme Action
    6. 6. Lock & Key Model <ul><li>Enzyme – Substrate Specificity! </li></ul>Substrate Enzyme Active Site
    7. 7. <ul><li>Usually large globular proteins (a) </li></ul><ul><li>b) Active Site where the substrate combines to the enzyme </li></ul><ul><li>c) Substrate which fits the active site </li></ul><ul><li>d) enzyme-substrate complex . The substrate is weakened to allow the reaction. </li></ul><ul><li>e) Unchanged enzyme/ re-used at low concentrations </li></ul><ul><li>f) Product of the reaction </li></ul><ul><li>Active site may have polar amino acids on the outside. </li></ul><ul><li>Enzyme specificity is due to the “complementary” shape of the active site and substrate. </li></ul><ul><li>Enzymes work at low concentrations. </li></ul>
    8. 8. The induced fit between an enzyme and its substrate
    9. 9. <ul><li>Active site is not a rigid pocket for the substrate to fit in. </li></ul><ul><li>Substrate “induces” the enzyme to change shape </li></ul><ul><li>Weakens the bonds of the substrate </li></ul><ul><li>Lowers the activation energy </li></ul>
    10. 10. The catalytic cycle of an enzyme
    11. 11. Inhibitors <ul><li>General function: </li></ul><ul><ul><li>Reduce activity of enzyme </li></ul></ul><ul><ul><li>Stop enzyme completely </li></ul></ul><ul><li>2 General types: </li></ul><ul><li>Competitive </li></ul><ul><li>Non Competitive </li></ul><ul><ul><ul><li>Irreversible </li></ul></ul></ul><ul><ul><ul><li>Allosteric – includes feedback inhibition </li></ul></ul></ul>
    12. 12. Metabolic Pathways <ul><li>Metabolic Pathways </li></ul>
    13. 15. Competitive Inhibitor <ul><li>Very similar to the substrate </li></ul><ul><li>Compete with the substrate on the active site </li></ul><ul><li>“stick” to the active site </li></ul><ul><li>NO reaction takes place </li></ul><ul><li>SLOW the rate of reaction </li></ul><ul><li>Prevents accumulation of substances </li></ul>
    14. 18. Competitive Inhibitor - Example <ul><li>Krebs Cycle </li></ul><ul><li>Malonate – inhibitor of cell respiration </li></ul><ul><li>Binds to the active site of the enzyme “Succinate dehydrogenase” </li></ul><ul><li>Interrupts Succinate to Fumarate step in the cycle </li></ul>
    15. 19. Krebs's Cycle – A METABOLIC PATHWAY
    16. 20. Non - Competitive <ul><li>Molecules NOT similar to the substrate </li></ul><ul><ul><li>NO competition for the active site </li></ul></ul><ul><li>“Bind” to another site on the enzyme </li></ul><ul><li>Inhibitor changes the conformation (3-D, tertiary structure) of the enzyme, causing enough alteration to slow enzyme activity; while substrate binds to the active site, it is not converted to a product. </li></ul>
    17. 22. TWO TYPES
    18. 23. Allosteric
    19. 24. Allsoteric <ul><li>FEEDBACK INHIBITION </li></ul>
    20. 25. Allosteric Inhibitors <ul><li>Non competitive (end – product inhibition) </li></ul><ul><li>Bind to a site in the enzyme: allosteric site . </li></ul><ul><li>RESULTS: </li></ul><ul><ul><li>Might change active site so substrate can fit </li></ul></ul><ul><ul><ul><ul><li>Increases reaction rate </li></ul></ul></ul></ul><ul><ul><li>Might “distort” active site and inhibit reaction </li></ul></ul><ul><ul><ul><ul><li>Lowers rate of reaction </li></ul></ul></ul></ul><ul><li>Can control the rate of metabolic reactions </li></ul><ul><ul><ul><li>ATP – feedback inhibition </li></ul></ul></ul>
    21. 27. Allosteric Inhibitors <ul><li>“feedback inhibition” </li></ul><ul><li>“ End product inhibition”: the last product of a reaction acting as an inhibitor for the another enzyme in the reaction. </li></ul>Metabolic Pathway Feedback Inhibition
    22. 28. Example <ul><li>ATP – Feedback inhibition (during glycolysis) </li></ul><ul><li>ATP accumulates </li></ul><ul><li>Acts as an allosteric inhibitor for the enzyme “phosphofructokinase.” PFK </li></ul><ul><li>Lowers the rate of reaction </li></ul><ul><li>Less ATP is produced </li></ul>
    23. 29. Irreversible inhibitors <ul><li>Generally are poisonous substances that enter from the outside of the body. </li></ul><ul><li>Binds to the enzyme “irreversibly” </li></ul><ul><li>Interferes with the binding of the enzyme with the substrate…metabolic pathway stopped </li></ul><ul><li>Examples: </li></ul><ul><ul><ul><li>CN (cyanide gas) – inhibits “cytochrome oxidase” during respiration </li></ul></ul></ul><ul><ul><ul><li>Lead and other heavy metals: binds to “sulfur” in enzymes </li></ul></ul></ul><ul><ul><ul><li>Nerve Gas – blocks an enzyme needed for the neurotransmitter acetylcholine </li></ul></ul></ul><ul><ul><ul><li>Antibiotics – penicillin – inhibits bacterial enzymes needed to build the Cell Wall – growth and reproduction STOPS </li></ul></ul></ul>
    24. 30. Inhibition Animations <ul><li>ACE Inhibitors </li></ul><ul><li>The Science of Enzyme Inhibition </li></ul>
    25. 31. Enzyme Action and Energy
    26. 32. How Do Enzymes Work? <ul><li>For a reaction to start it needs energy: </li></ul><ul><li>ACTIVATION ENERGY </li></ul><ul><li>Enzymes LOWER the Activation Energy </li></ul>
    27. 33. Energy Progress of Rxn. Reactants Products
    28. 34. Energy Progress of Rxn. Reactants Products Activation Energy - Minimum energy to make the reaction happen
    29. 35. Energy Progress of Rxn. Reactants Products Enzyme Substrate Complex or Transition State
    30. 36. Energy Progress of Rxn. Reactants Products Overall energy change
    31. 37. Energy Progress of Rxn. Reactants Products Exergonic
    32. 38. Enzymes and Reactions <ul><li>Reactions are not impossible without enzymes. </li></ul><ul><ul><li>With enzymes the RATE of reactions will increase. </li></ul></ul><ul><ul><li>Enzymes do no change the “contents” of the reactions. </li></ul></ul>
    33. 39. Energy in a Reaction <ul><li>Reactants must absorb energy from their surroundings for their bonds to break. </li></ul><ul><li>Products release energy when their new bonds are formed. </li></ul><ul><li>The amount of energy needed for a chemical reaction to proceed is the ACTIVATION ENERGY, E A. </li></ul>
    34. 40. Endergonic vs. Exergonic
    35. 41. Endergonic Reaction <ul><li>Endergonic Reaction (Endothermic) – </li></ul><ul><ul><li>Absorb energy into the reaction </li></ul></ul><ul><ul><li>Amount of energy in system increases </li></ul></ul>
    36. 42. Endergonic Rxn.
    37. 43. Exergonic Reaction <ul><li>Give off Energy </li></ul><ul><ul><li>Represented by a negative change in energy </li></ul></ul><ul><ul><li>Loss of free energy </li></ul></ul>
    38. 44. Energy profile of an exergonic reaction
    39. 45. Figure 6.13 Enzymes lower the barrier of activation energy
    40. 46. Catalyzed Reactions <ul><li>The Small Intestine Reactions: </li></ul><ul><li>Starch + Amylase = many maltose units </li></ul><ul><li>Maltose + maltase = glucose + glucose </li></ul><ul><li>Lactose + lactase = glucose + galactose </li></ul><ul><li>Sucrose + sucrase = glucose + fructose </li></ul><ul><li>These are ALL hydrolytic reactions!! </li></ul>
    41. 47. A Cell Reaction <ul><li>Cytotoxic Reaction: </li></ul><ul><li>2H 2 O 2 + catalase  O 2 + 2H 2 O </li></ul><ul><li>Gas Exchange: CO 2 transport in blood </li></ul><ul><li>CO 2 + H 2 O + carbonic anhydrase  H 2 CO 3 </li></ul>
    42. 48. Figure 6.15 The catalytic cycle of an enzyme
    43. 49. Characteristics of Enzyme Activity <ul><li>1. Enzymes work best at certain temperatures and enable cell reactions to proceed at normal temperatures. </li></ul><ul><li>2. Small amount of enzyme can affect large amounts of substrate. </li></ul><ul><li>3. Enzyme and Substrate concentration can control the rate of the reaction. </li></ul><ul><li>4. Enzymes work best at a certain pH. </li></ul><ul><li>5. A Co-enzymes may be needed for an enzyme to function correctly. </li></ul>
    44. 50. Characteristics of Enzyme Activity <ul><li>THE MAD GOOD ANNY’ </li></ul>
    45. 51. Figure 6.16 Environmental factors affecting enzyme activity
    46. 52. Temperature <ul><li>As you increase temperature , enzyme action increases as well until an optimum temperature for enzyme action is reached. </li></ul><ul><li>Often doubling with every 10 °C rise </li></ul><ul><li>“ collisions” between substrate and active site more frequent at higher temps. </li></ul>
    47. 53. Enzymes are temperature dependent…... (in humans) <ul><li>Most work at body temperature:37 o C to maintain homeostasis </li></ul><ul><li>Denature at high temperatures </li></ul><ul><li>Inactive at low temperatures </li></ul>
    48. 54. Effect of Substrate Concentration <ul><li>As substrate concentration increases, the rate of enzyme catalyzed reactions will increase….and then become constant . </li></ul><ul><li>The increased concentration is a “limiting factor” </li></ul><ul><li>When all active sites are engaged by substrates, reaction rate will not increase. </li></ul>
    49. 55. pH <ul><li>Affects enzyme action. </li></ul><ul><li>Certain enzymes work better in acidic environments while others in basic environments. </li></ul>
    50. 56. Effect of pH on Enzyme Activity <ul><li>Enzymes have an optimum pH at which they function best. </li></ul><ul><li>At other pH’s, enzymes are denatured , because the H + and OH - ions disrupt hydrogen bonds that hold the 3-D structure in place. </li></ul>Effect of pH on two different enzymes: pepsin and trypsin
    51. 57. Enzymes are pH specific. <ul><li>Different enzymes </li></ul><ul><li>Different body areas </li></ul><ul><li>Different optimum pH </li></ul><ul><li>Examples: </li></ul><ul><li>Stomach= 1.5 to 4.0 </li></ul><ul><li>Mouth = 6.5 – 7.5 </li></ul><ul><li>Blood = normal – 7.4 </li></ul>Blood
    52. 59. Co-existing with Enzymes: <ul><li>Cofactors and Coenzymes </li></ul>
    53. 60. Cofactors <ul><li>Needs to be present in addition to an enzyme for a reaction to catalyze. </li></ul><ul><li>Like enzymes, they return to their original state when their reactions are completed. </li></ul><ul><li>Inorganic – potassium, zinc, magnesium, iron (not H 2 O) </li></ul>
    54. 61. COENZYMES <ul><li>A type of cofactor ---- ORGANIC in nature </li></ul><ul><ul><ul><li>Separate from the protein in the enzyme to react DIRECTLY in the chemical reaction. </li></ul></ul></ul><ul><ul><ul><li>Function: transfer electrons, atoms, or molecules between enzymes </li></ul></ul></ul><ul><li>Vitamins – help make coenzymes </li></ul><ul><ul><ul><li>Vitamin B – helps to make NAD a coenzyme in cellular respiration. </li></ul></ul></ul><ul><ul><ul><li>NAD – electron carrier </li></ul></ul></ul>
    55. 62. COENZYMES
    56. 63. Application of Enzymes in Biotechnology
    57. 64. <ul><li>Pectinase – ******IB Req. </li></ul><ul><li>Pectin – polysaccharide found in fruit skins (cell wall) </li></ul><ul><li>Pectinase – found in natural fungus (Aspergillus niger) on fruit. Uses enzyme to soften fruit. </li></ul><ul><li>Hydrolyzes pectin into its monomers. </li></ul><ul><li>Pectinase is added during the crushing of fruit </li></ul><ul><li>Makes juice clearer & juicier! Easy to separate from pulp. </li></ul>Making Juice
    58. 65. Lactose intolerance <ul><li>Unable to produce </li></ul><ul><li>lactase in sufficient </li></ul><ul><li>quantities. </li></ul><ul><li>Usually treated with </li></ul><ul><li>supplements OR lactose </li></ul><ul><li>free products </li></ul>
    59. 66. <ul><li>Biological Detergents </li></ul><ul><li>Proteases – protein (most dirt) </li></ul><ul><li>Source: Bacteria – B acillus licheniformis </li></ul><ul><li>Contain enzymes to digest stains at lower temperatures than 45 C and high pH’s </li></ul><ul><li>Wash in cold water!...lower energy use, less shrinkage. </li></ul>
    60. 67. Making Cheese <ul><li>Rennin – an enzyme that turns milk to cheese. </li></ul><ul><li>Coagulates into curd at body temperature </li></ul><ul><li>Found in stomachs of babies – helps to retain food </li></ul><ul><li>Found in stomachs of young cattle***** </li></ul>
    61. 68. Milk <ul><li>Adding LACTASE to milk. </li></ul><ul><li>Breaks down LACTOSE into easier digestible sugars </li></ul><ul><li>Removes over 70% of lactose </li></ul><ul><li>Prevents painful stomach disorders….in children BUT… </li></ul><ul><li>More expensive </li></ul><ul><li>May lose ability to make lactase </li></ul>

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