Isoenzymes, also known as isozymes, are different forms of the same enzyme that catalyze identical chemical reactions but vary in properties such as electrophoretic mobility and tissue localization. Lactate dehydrogenase (LDH) has multiple isoenzymes with clinical significance in diagnoses like myocardial infarction and various diseases affecting the kidneys and muscles. Creatine phosphokinase (CK) also has isoenzymes important for assessing brain and muscle injuries, while alkaline phosphatase (ALP) isoenzymes are significant in liver and bone disorders.

1. Isoenzymes
Clinical enzymology

2. 2
ISOENZYMES
 Isoenzymes or isozymes are mutiple
forms of same enzyme that catalyse the
same chemical reaction
 Different chemical and physical properties:
 Electrophoretic mobility
 Kinetic properties
 Amino acid sequence
 Amino acid composition

3. 3
S. No Property E.g.
1 Electrophoretic
mobility
Isoenzymes of Lactate dehydrogenase have
different electrophoretic mobility
2 Heat stability Alkaline phosphatase isoenzymes are either
heat labile or stable
3 Inhibitor An inhibitor can inhibit only one isoenzyme of
an enzyme eg.Acid phosphatase
4 Km Glucokinase and hexokinase
5 Cofactors Mitochondrial isocitrate dehydrogenase requires
NAD+
, cytosolic form requires NADP+
6 Tissue localisation LDH 1 is present in heart, LDH 5 in muscle
7 Antibodies For creatine kinase, each isoenzyme can be
bound only by a specific antibody

4. 4
Lactate dehydrogenase (LDH)
 E.C – 1.1.1.27
 L-lactate :NAD+
oxidoreductase:LDH
 Molecular weight- 134 kDa
 tetramer
 M (A) -muscle –chromosome 11(basic)
 H (B) -heart – chromosome 12(acidic)

5. 5
Lactate dehydrogenase (LDH)
Normal values
Serum -100 -200 U/L
CSF - 7 -30 U/L
Urine - 40 -100 U/L

6. 6
Isoenzyme
name
Composition Electrophore
tic migration
Present in Elevated in
LDH 1
Heat
resistant
( H4) Fastest
moving
Myocardium,
RBC,kidney
myocardial
infarction
LDH2
Heat
resistant
(H3M1) Myocardium,
RBC,kidney
Kidney
disease,megalo
blastic anemia
LDH3 (H2M2) brain Leukemia,malig
nancy
LDH4
Heat labile
(H1M3) Lung,spleen Pulmonary
infarction
LDH5
Heat labile
Inhibited by
urea
(M4) Slowest
moving
Skeletal
muscle, Liver
Skeletal muscle
and liver
diseases

7. 7
Lactate dehydrogenase (LDH)
This is an example in which two duplicated genes have
become specialized to different tissues.
The isozymes are also differentially expressed in different
developmental stages. Before birth the heart is more
anaerobic compared with adulthood. Indeed, before birth
the main isozyme in the heart is the M4, and with time it
switches to HM3 (at birth), to H2M2 and HM3 at 1 year
after birth, and to H3M AND H4 after 2 years.
My main LDH is HM3
My main LDH is HM3. Great!

8. 8
Atypical forms of LDH
 sixth isoenzyme LDH- X
 Seventh isoenzyme – LDH -6

9. 9
Clinical significance of LDH
 Myocardial infarction (LDH 1>LDH2)
 Megaloblastic anemia (50 times upper limit of
LDH 1 and LDH 2)
 Muscular dystrophy (LDH 5)
 Toxic hepatitis with jaundice (10 times more
LDH 5)
 Renal disease- tubular necrosis or
pyelonepheritis
 Pulmonary embolism LDH 3 (massive
destruction of platelets)

10. 10
 Leukemia (LDH 2 and LDH 3)
 Malignancy (LDH 3)
 Hodgkins disease
 germ cell tumors
 Urinary LDH-3 to 6 times normal:
chronic glomerulonephritis
Systemic lupus erythematosus
Diabetic nephrosclerosis
Bladder and kidney malignancies

11. 11
 In CSF:
 Bacterial meningitis – LDH 4 and LDH 5
 Viral meningitis - LDH 1
 Metastatic tumors - LDH 5
 Neonatal cases of intracranial
haemorrhage associated with seizures and
hydrocephalus

12. 12
LDH in starch gel
The H(B) monomer
is very negatively charged

13. 13
CREATINE PHOSPHOKINASE
 Adenosine triphosphate:creatine N-
phosphotransferase
 E.C-2.7.3.2
 Dimeric enzyme (82 kDa)
 4 -60 IU/L

14. 14

15. 15
 Enzyme unstable in serum
 Activity lost due to sulfhydryl group
oxidation at active site
 Dimer (each of 41000 Da)
 B (brain) – chromosome 14
 M (muscle) –chromosome 19

16. 16
Isoenzy
me
name
Compo
sition
Present in Elevated in
CK-1
Fast
moving
BB
Brain,prostate,GI
tract,lung,bladder,uteru
s,placenta
CNS diseases
CK-2
2% of
total
MB Myocardium/ Heart
Acute myocardial
infarction
CK-3
Slow
moving
MM
Skeletal muscle,
Myocardium
All 3 in cytosol

17. 17

18. 18
atypical forms of CK
 Fourth form - CK-Mt (chromosome 15)
severe illness
Malignant tumors
 macroCK
 type 1- CK BB complexed with IgG
 type 2-oligomeric CK-Mt

19. 19
Clinical significance of CK
 CK 1 elevated:
very low birth weight newborns
brain damage in neonates
neurological injury –CK 1 rise in CSF
>200 U/L –die
100 – 200 U/L – survive with
neurological defecits
<100 U/L – good chance of
recovery

20. 20
Elevated CK 1
 Adenocarcinomas of GI tract
 Carcinoma lung
 Ca
prostate,bladder,testes,kidneys,breast,ova
ries,uterus,CNS,leukemia,lymphoma and
sarcoma

21. 21
 A 66-year-old man sought medical care at
the hospital due to severe chest pain
lasting for 24 hours.The patient was
aware of being hypertensive and was a
smoker.Without any prior symptom, he
started to have severe chest pain and
sought emergency medical care after
about 24 hours, due to pain persistence.

22. 22
ECG- low QRS voltage in the frontal plane, electrical alternan of QRS
complexes, electrically inactive lower wall area and extensive ongoing
myocardial infarction.

23. 23
 Elevated CK 2:
myocardial infarction
head injuries
subarachnoid haemorrhage
exercise
Elevated CK 3:
muscular dystrophies(DMD- 10000 IU/L)
myopathies
hypothyroidism (5 fold more than normal
value,also CK 2 is elevated)

24. 24
Alkaline phosphatase (ALP)
 E.C -3.1.3.1.
 Orthophosphoric monoester
phosphohydrolase
 In mucosa of small intestine, proximal
convoluted tubule, bone, liver, placenta
 Catalyses alkaline hydrolysis of naturally
occuring and synthetic substrates

25. 25
Isoenzymes of ALP
 Alpha 1 ALP-epithelial cells of biliary canaliculi
 Alpha 2 heat labile ALP- hepatic cells
 Alpha 2 heat stable ALP-not destroyed at 65˚C
 inhibited by phenylalanine
 placental
 Pre beta ALP – bone,heat labile
 Gamma ALP – intestinal cells
 inhibited by phenylalanine
 Leukocyte alkaline phosphatase –decreased in CML
 increase in lymphoma
ATYPICAL ISOENZYMES
 Regan isoenzyme-heat stable,inhibited by L-phenylalanine
 Nagao isoenzyme- variant of regan
 inhibited by L-leucine

26. 26
Clinical significance
 Hepatobiliary disease
 Hepatic carcinoma
 Hepatic metastases
 Pagets disease (10 – 25 times)
 Bone cancer
 Healing of bone fracture
 Osteomalacia and rickets
 Hyperparathyroidism
 Ca of ovary,uterus-regan isoenzyme
 Metastatic Ca of pleural surfaces –Nagao isoenzyme