Enzymes - fast, specific catalysts

What are enzymes?

• All enzymes are globular proteins → spherical in shape (Fig 1)
• Control biochemical reactions in cells
• They have the suffix "-ase"
• Intracellular enzymes are found inside the cell
• Extracellular enzymes act outside the cell (e.g. digestive enzymes)
• Enzymes are catalysts → speed up chemical reactions (Fig 2, Fig 3)
  • Reduce activation energy required to start a reaction between molecules
  • Substrates (reactants) are converted into products
  • Reaction may not take place in absence of enzymes (each enzyme has a specific catalytic action)
  • Enzymes catalyse a reaction at max. rate at an optimum state
    
• Lock and key theory
  • Only one substrate (key) can fit into the enzyme's active site (lock)
  • Both structures have a unique shape
• Induced fit theory (Fig 4)
  • Substrate binds to the enzyme's active site
    • The shape of the active site changes and moves the substrate closer to the enzyme
    • Amino acids are moulded into a precise form
    • Enzyme wraps around substrate to distort it
  • This lowers the activation energy
  • An enzyme-substrate complex forms → fast reaction
  • E + S → ES → P + E
    
• Enzyme is not used up in the reaction (unlike substrates)

Fig4

Fig1

Enzyme Activity

• Changes in pH
  • Affect attraction between substrate and enzyme
  • Ionic bonds can break and change shape → enzyme is denatured
  • Charges on amino acids can change → ES complex cannot form
  • Optimum pH (enzymes work best)
    • pH 7 for intracellular enzymes
    • Acidic range (pH 1-6) in the stomach for digestive enzymes (pepsin)
    • Alkaline range (pH 8-14) in oral cavities (amylase)
  • pH measures the conc. of hydrogen ions → higher conc. will give a lower pH
• Enzyme conc
  • Proportional to rate of reaction, provided other conditions are constant
  • Straight line
• Substrate conc. (Fig 5)
  • Proportional to rate of reaction until there are more substrates than enzymes present
  • Rate of reaction increases
    • Substrate binds to active site, but more enzymes are available
    • Rate increases if more substrate is added
  • Eventually, curve becomes constant (no increased rate)
    • Substrates occupy all active sites (all enzymes)
    • Adding more substrate won't yield more product, as no more active sites are available

• Increased Temperature
  • Increases speed of molecular movement → chances of molecular collisions → more ES complexes
  • At 0-42°C rate of reaction is proportional to temp
  • Enzymes have optimum temp. for their action (usually 37°C in humans)
  • Above ≈42°C, enzyme is denatured due to heavy vibration that breaks -H bonds
  • Shape is changed → active site can't be used anymore
• Decreased Temperature
  • Enzymes become less and less active, due to reductions in speed of molecular movement
  • Below freezing point
    • Inactivated, not denatured
    • Regain their function when returning to normal temperature
  • Thermophilic: heat-loving
  • Hyperthermophilic: organisms are not able to grow below +70°C
  • Psychrophiles: cold-loving

Enzymes - Heroes and Villains

Analytical reagents

• Made up of 2 enzymes (glucose oxidase and peroxidise) and a colourless hydrogen-donor fixed on a strip
  • The strip is dipped into a test solution (urine)
  • Colour develops which indicates that glucose is present
  • This method is used by diabetics to monitor their blood glucose levels
  • In healthy people, the urine contains NO glucose
• Glucose oxidase
  • Highly sensitive to low conc. of glucose
  • Highly specific because it only reacts with one specific substrate (glucose)
  • Catalyses the conversion of glucose to hydrogen peroxide (H2O2)
• Peroxidase
  • Catalyzes reaction between colourless hydrogen-donor molecule and H2O2
  • A coloured molecule is formed

Alpha1-antitrypsin

• Function
  • White blood cells (neutrophils) in the lung help to prevent infections
  • They also release elastase and protease (trypsin)
  • Those enzymes break down/digest ct. and proteins inside the lungs and damage it
  • NB: Trypsin is also found in the digestive system and digests food!
  • The anti-protease alpha1-antitrypsin protects the lungs from elastase and protease
• Alpha1-antitrypsin deficiency
  • Genetic disease that causes emphysema
  • Trypsin is no longer inhibited and damages the lungs
  • Walls of alveoli are damaged and surface area for gas exchange is reduced
  • Patients can be treated by infusing alpha1-antitrypsin
• Smoking
  • Increases the number of neutrophils in the lungs (more trypsin is secreted into the lungs)
  • ALSO inactivates alpha1-antitrypsin
  • This creates an imbalance between proteases (trypsin) and anti-proteases (α1-antitrypsin)
  • Same lung damage as in α1-antitrypsin deficiency but much slower

Lactose intolerance

• Lactase splits lactose (milk sugar) into β-glucose and galactose
• Lactose intolerant person lacks lactase → lactose is neither digested nor absorbed
• High levels of soluble lactose remain in small intestine
• Supports large populations of bacteria / ferment lactose producing gas / causing abdominal discomfort
• Water potential becomes more negative / H2O moves into small intestine / not reabsorbed / diarrhoea
• Adults rarely produce lactase / gene is switched off in adulthood

Pancreatitis

• Pancreas is found below the stomach
• Produces digestive enzymes (amylase, lipase, trypsin that break down starch, lipids, proteins, respectively)
• Acute (sudden onset)
  • Reversible inflammation of a previously normal pancreas
  • Caused by gallstones, alcohol, scorpion bite, trauma
  • Inappropriate activation of enzymes
    • Trypsin becomes active before released from the pancreas
    • Pancreas is made of proteins
    • Trypsin is active and digests/hydrolyses proteins
    • Cell wall breaks down, amylase and lipase escape into the blood
  • Diagnosis
    • Amylase and lipase remain elevated in blood for 3 days
• Chronic (gradual onset)
  • Inflammation is caused by cystic fibrosis or long-term alcohol intake
  • Pancreas gradually loses its ability to produce digestive enzymes
  • Food is not digested, hence not absorbed (malabsorption)
  • Diagnosis
    • Amylase and lipase in blood normal
    • Low levels of faecal elastase
    • Too much fat in faeces → fat passes through gut without being digested and absorbed
• Pancreatic enzyme replacement therapy (PERT)
  • Required for life in patients with irreversible pancreatic damage
    • Chronic pancreatitis
    • Cystic fibrosis
  • Tablet contains digestive enzymes (normally produced by pancreas)
  • Degradation of enzymes/proteins by stomach acid prevented by
    • Coating tablet with protective layer
    • Taking extra tablet that inhibits acid secretion