1. Enzymes-1 – Structure and Isoenzymes lecture NO : 1st MBBs
Dr Muhammad Ramzan

2. Enzyme (E) – the definition
A protein that catalyzes chemical reactions of other substances
without itself being destroyed or altered upon the:
Completion of the reactions.
Enzymes lower the activation energy
Enzymes are produced by the living organisms

3. Enzymes (E) – the background Intra/extra cellular
All enzymes are proteins except a few catalytic RNAs- Ribozymes
Defect or deficiency of an enzyme produces diseases called "inborn error of metabolism.“ like Lactose Intolerance
The human body contains about 10,000 different enzymes
Some enzymes work within the cell ( RNA polymerase) and some outside the cells like digestive enzymes
The substance acted upon an by E is called a Substrate - S

4. Lactose intolerance

5. Biological significance of Enzymes Reduces activation energy
At body temperature, very few biochemical reactions proceed at a significant rate without the presence of an E
Enzymes lower the activation energy of a catalyzed reaction 2
Thus increasing the rate of the reaction
They do so in a step by step, highly efficient and safe manner 3
Enzymes uses activation energy which is the minimum energy required to start and complete a reaction

6. Enzyme lowers the activation energy

7. Properties of Enzymes
Catalysis: act as protein catalysts - ↑ the velocity of reaction
Activation energy: is reduced- energy to start and complete a reaction
Active site binds and catalyze the S via side chains of its AAs
Catalytic efficiency: E are efficient - Turn over no: is usually 103 – 108
Specificity: E is specific for specific S – Catalyzes 1 type of reaction
Regulation: E activity is regulated by cell according to body needs
Location of E- Compartmentalization – intra/extracellular

8. Parts of Enzymes(E) - 2 parts body and active site
Enzymes are macromolecular proteins that are commonly globular in shape and variable in size -
Each Enzyme has a specific area to bind the substrate by non covalently/temporary
Essentially enzymes have 2 parts:
Body of enzyme and
Active site

9. Structure of enzyme – 2 parts Body and active site

10. Structure of enzymes – the body 1 Variable in size
Enzymes are the globular proteins and their AA content vary from 62 to 2500 AA residues ( Fatty acid Synthase. FAS )
E are larger than S and some are grouped together to form enzyme complex like FAS/LDH
Enzymes are long linear chains of AAs but their activities are due to their 3D Tertiary structures
Apozyme is the protein portion (inactive E) while Halozyme is a protein with a cofactor/coenzyme (Active E)

11. Structure of enzyme Apozyme and Halonzyme

12. Structure of enzymes – the body

13. Structure of E – the active site 2 produced by protein folding
Active sites are the hollows/cracks/ pockets on the surface of E and are produced by protein folding- 3D structure
Active site contains 2-4 AA that are involved directly in S binding and catalysis (+ electrical+ chemical properties of AA side chains)
This binding is temporary by non covalent bonds and is relieved after the completion of the catalysis
S of just right shape and attractive groups/sequence of AAs can fit into these active sites
The usual analogy for this is a key fitting exactly into the lock

14. Active site – the chemistry

15. Enzymes - Principle of working 1 E identifies and binds S
Enzyme identifies the Substrate (S) and works by binding the Substrates so that they(S) are held in a :
Particular Geometric configuration
Forms Enzyme Substrate Complex (ESC) by binding E+S
The binding is temporary (non covalent),
E lowers the activation energy which is essential for the reaction to occur and complete

16. Enzymes – Principal of working 2 E controls the direction of reaction
Both, the enzyme and products are released upon completion of the reaction and enzyme is available for Reuse
Like other catalysts, an enzyme does not control the direction of the reaction. It is controlled by the cell itself
Cell ↑the rates of forward and reverse reactions proportionally depending upon the body needs (Regulation)

17. Working of enzymes – the principals

18. Enzymes – Quantity and location Within and outside the cell
Most enzymes are produced in tiny quantities and catalyze reactions that take place within the cells like .
DNA/ RNA Polymerase and HMG CoA Synthase/Catalase
Digestive enzymes, however, are produced in relatively large quantities and act outside the cells in the lumen of the GIT
They cleave the dietary macromolecules like TG,Glycogen and Nucleic acids into smaller ones and monomers

19. Allosteric or Additional sites Allosteric enzymes
Enzymes can also have sites other than active site that bind cofactors, which are needed for catalysis.(multi units)
Some E also have binding sites for small molecules, which are often direct/ indirect products /S of the reaction catalyzed.
This binding can serve to ↑or↓ (regulate) the enzyme's activity, providing a means for feedback regulation.
They are the Allosteric sites and enzyme as Allosteric enzymes
Glycogen Phosphorylase- that breaks down the glycogen when glucose is needed

21. Allosteric regulation

22. Allosteric sites and enzymes

23. Denaturation of enzymes – heat and pH
Most enzymes can be denatured - unfolded and inactivated
by heating or chemical denaturants / change of pH
It disrupts the 3 structure of the protein (E) and results in the loss of function
Denaturation of E may be reversible or irreversible depending upon the damage to the enzyme

24. E. Denaturation – high temperature

25. E denaturation- change of pH Reversible Denaturation

26. Isoenzymes/ Isozymes – the Definition difference in sequence of AAs
Isoenzymes are generally the Multiple forms of the enzymes that differ in AA sequence but catalyze the :
Same chemical reaction.
These include Lactic Dehyrogenase (LDH) ,Creatine Kinase (CK) and Alkaline Phosphatase

27. Isoenzymes - Characteristics
These display different electrophoretic mobility, regulatory and immunological properties
Isozymes are the result of gene duplication
These genes express the polypeptides chains that produce different subunits of the Isoenzymes
Like CK has 2 Polypeptide subunits – M and B
These include Lactic Dehydrogenase (LDH) ,Creatine Kinase (CK) and Alkaline Phosphatase

28. Isoenzymes – the clinical significance
Different organs of the body contain the characteristic proportions of the different Isoenzymes
The pattern of Isozymes in the plasma may identify the site of tissue damage like :
Creatine Kinase has 3 forms and is present in various tissues
The damage to these tissues releases  amount of respective CK Isoenzymes in circulation (Heart, brain and muscles)

29. Isoenzymes of CK (3) – the associated diseases
There are 3 Isoenzymes of the CK including:
CKMM In skeletal muscle diseases.
CKMB cardiovascular diseases (MI)
CKBB in brain and intestinal diseases like infarction

30. Creatine Kinase – Clinical importance
Level of Creatine Kinase CK (CKMB) is commonly determined for the diagnosis of Myocardial infarction – MI
Isoenzymes are particularly useful when ECG is difficult to interpret especially when there have been :
Previous episodes of MI
Same is true for Cardiac Troponin T and I