2 biochemistry and metabolism

744 views
555 views

Published on

dr ehab

Published in: Education
0 Comments
2 Likes
Statistics
Notes
  • Be the first to comment

No Downloads
Views
Total views
744
On SlideShare
0
From Embeds
0
Number of Embeds
1
Actions
Shares
0
Downloads
26
Comments
0
Likes
2
Embeds 0
No embeds

No notes for slide

2 biochemistry and metabolism

  1. 1. Biochemistry and MetabolismThe course will cover:Enzymes – kinetics and controlControl of metabolismControl of metabolism is the control of enzyme activity
  2. 2. Biochemistry and MetabolismWhy should we study metabolism?Why is metabolism interesting?Where does metabolism feature in daily life?Why should we study enzymes?Why are enzymes interesting?How do enzymes feature in daily life?
  3. 3. from Drug Discovery Today 11 481-493 (2006)
  4. 4. ―statins‖Inhibitors of the enzyme HMG CoA reductase
  5. 5. HMG CoA reductaseHMG CoA + NADPH → mevalonate + NADPRate limiting, regulatory enzymein cholesterol synthesis
  6. 6. Cholesterol and heart diseaseCholesterol as key factorExcessive deposition in blood vessels leads to heart diseaseCarried as lipoproteins“bad” cholesterol – low density lipoprotein (LDL)“good” cholesterol – high density lipoprotein (HDL)
  7. 7. Benefits of LipitorLowers total cholesterol by ~ 40%.Lowers "bad," low-density lipoprotein (LDL) cholesterol by ~ 50%.Reduces risk for heart attack and stroke
  8. 8. Every five minutes somebody in the UK dies from a heart attack
  9. 9. Deaths by cause, women, 2004, United Kingdom Injuries & poisoning Respiratory disease 3% 14% All other causes Other cancer 22% 14% Colo-rectal cancer 2% Breast cancer 4% Lung cancer 4% Coronary heart disease 15% Other CVD 9% Stroke 12%Office of National Statistics (2005)Scotland General Register Office (2005)Northern Ireland General Register Office (2005) www.heartstats.org
  10. 10. Risk factors for heart diseaseHigh fat dietOverweight/obeseHigh blood pressureSmokingLack of exerciseHeredity
  11. 11. Increase in obesity
  12. 12. Prevalence of overweight and obesity by age, women, latest available year, England, Scotland and Wales 16-24 80 25-34 35-44 70 45-54 55-64 65-74 60 75 & over Overweight/obesity (%) 50 40 30 20 10 0 England (2004) Scotland (2003) Wales (2003/04)Overweight/obesity: BMI 25 kg/m2 and overHealth Survey for England 2004; Scottish Health Survey 2003:Welsh Health Survey 2003/04 www.heartstats.org
  13. 13. lifestyleless more
  14. 14. diet Guideline daily amountFood Standards Agencytraffic light labelling
  15. 15. Metabolic syndrome1. Obesity, particularly around the waist (having an "apple shape")2. Elevated blood pressure3. An elevated level of triglycerides and a low levelof high-density lipoprotein (HDL) — the "good" cholesterol4. Resistance to insulin
  16. 16. Metabolic syndromePrevalence – 20% of adults in USRisk Factors: poor eating habits,lack of exercise, genetics, ageing
  17. 17. Metabolic syndrome1. Increased risk of heart disease2. Increased risk of Type 2 diabetes
  18. 18. Questions to ask about diet and metabolismHow does a high carbohydrate diet lead to obesity?How does a high fat diet lead to heart disease?What is disturbed in diabetes?
  19. 19. Complex interplay between glucose/fat/cholesterol metabolism
  20. 20. Need to understand diet and metabolismDaily Mail:August 20th 2002 – Could this be the cure for obesity?July 30th 2003 – Chemical could burn away obesityGillian McKeith – “she hasnt a clue about nutrition” (Brit Dietetic Assn)
  21. 21. Biochemistry and MetabolismWhy should we study enzymes?Why are enzymes interesting?How do enzymes feature in daily life?
  22. 22. Enzymes?Biological washing powdersDrug targetsTests for disease/test kitsClinical tests for liver damage
  23. 23. Biological washing powdersContain lipases and proteases to digest fat and protein in stainsWork at lower temperatures
  24. 24. Drug targetsInhibitors of enzyme – COX-2 (cyclooxygenase-2,responsible for prostaglandin synthesis)
  25. 25. ELISAEnzyme Linked ImmunoSorbant AssaysUsed widely for detection of proteins and antibodies in patient samplesUses enzyme conjugated to antibody
  26. 26. Test for HIVPatients with HIV have antibodies against the viral proteinsDetect these antibodies using ELISA
  27. 27. 96 well plate for ELISA
  28. 28. ELISA for antibodies in HIVEnzyme-labelled Colour development afterAnti-human antibody adding substrate for enzymePatient sample –Containing anti HIV HIV proteins
  29. 29. ELISADetection antibody has enzyme attached to ite.g. alkaline phosphatase – substrate 4-nitrophenyl phosphate NO2 NO2 O + HO P O O OH HO P O O OH colourless yellow
  30. 30. 96 well plate for ELISA
  31. 31. Pregnancy test kits
  32. 32. Tests for liver functionDoctor requests “liver enzymes”Blood testMeasure total protein, albumin and several enzymes in serum
  33. 33. Liver enzymesAlanine aminotransferaseAspartate aminotransferaseEnzymes involved in amino acid metabolism
  34. 34. Liver enzymes in blood sampleAlanine aminotransferase Leaks in to blood from damaged liver cells e.g. viral hepatitis, paracetamol overdose, fatty liver (alcohol abuse) etcAspartate aminotransferase Also raised in liver damage but not as specific
  35. 35. What do we need to know about enzymes?How to set up an enzyme assayEffects of inhibitorsHow to interpret the results
  36. 36. HMG CoA reductase assayHMG CoA + NADPH → mevalonate + NADPAssay from loss of absorbance at 340 nmas NADPH consumedMix enzyme and substrate in cuvette and read A340
  37. 37. Kinetics of enzymes 6 5NADPH (A340) ―initial rate‖ 4 3 2 1 0 0.0 2.5 5.0 7.5 10.0 12.5 time
  38. 38. Effect of statin on HMG CoA reductase activity 25 control 20 15 + statin rate 10 5 0 0.000 0.025 0.050 0.075 0.100 [substrate]
  39. 39. Effect of statin on HMG CoA reductase activityCompetitive inhibition from FEBS Letts 72 323-326 (1976)
  40. 40. Enzyme assays in “real life”Hospital pathology lab Industrial drug discovery

×