Enzymes

• Catalyzes reaction
  

DEFINITION OF TERMS

• Apoenzyme: protein portion of enzyme without cofactor
  
• Holoenzyme: complete active enzyme
  
• Active site: site where enzymatic reaction occurs
  
• Allosteric site: site other than the active site
  
• Isoenzyme: forms of enzyme that are different from each other but still catalyzes same reaction
  

CATEGORIES

• Oxidoreductase
  
  • For oxidation/reduction reactions
    
  • Ex: LDH, G6PD and Malate dehydrogenase
    
• 2. Transferase
  
  • Catalyzes transfer of groups from one substrate unto another
    
  • Ex: AST, ALT, CK, GGT
    
• Hydrolase
  
  • Hydrolysis
    
  • Ex: ACP, ALP, 5’NT, AMS, LPS, CHS
    
• Lyase
  
  • Removal of groups but with no hydrolysis
    
  • Ex: Aldolase
• Isomerase
  
  • Interconversion of isomers
• Ligase
  
  • Joins to 2 substrate molecules
  • Ex: synthases

HEPATIC ENZYME PROFILE

1. ALP

• Sources
  • Liver
  • Kidney
  • Bone
  • Placenta
  • Intestine
  • WBC

• Methods	Substrates
  Bodansky Shenowara Jones King-Armstrong Bessy Lowry-Brock	Β-glyceroPO4 Β-glyceroPO4 p-nitrophenylPO4 p-nitrophenylPO4

• Facts
  • Optimum pH: 10
  • Greatly elevated in Paget’s disease
  • Avoid using EDTA-CitrateOxalate

2. ALT (SGPT)

• Sources
  • Liver
  • RBCs

• Methods	Substrates
  Reitman-Frankel (DNPH)	Alanine α-keto

• Facts
  • Liver-specific
  • Marked elevation with viral hepatitis
  • De ritis ratio:
    • >1 = viral;
    • <1 = non-viral

3. LD

• Sources
  • All Tissues
• Methods
  • Wacker Method (forward)
  • Wrobleuski La Due (reverse)
  • Wrobleuski Cabaud
  • Berger Broida
• Facts
  • NAD+ (cofactor)
  • LD4 and LD5  Storage: 25°C upto 24 hours

4. GGT

• Sources
  • Canaliculi of hepatic cells
  • Kidney
  • Prostate 
  • Pancreas

• Methods	Substrate
  SZAZ	Gammaglutamylp-nitroanilide

• Facts
  • Most sensitive marker for alcoholic hepatitis

CARDIAC ENZYME PROFILE

1. CK

• Sources
  • Cardiac
  • skeletal 
  • brain tissues
• Methods
  • Tanzer-Gilvarg (forward)
  • Oliver-Rosalki (reverse)
• Facts
  • CK-BB (fastest migrating; most anodal),
  • CK-MB, CK MM (slowest; least anodal)
  • Sensitive indicator of AMI & Duchenne disorder
  • Highest elevation of total CK: Duchenne’s muscular dystrophy
  • Light and pH sensitive

2. AST (SGOT)

• Sources 
  • Liver
  • heart
  • skeletal muscle

• Methods	Substrate
  Karmen Method (Ph 7.5; 340 nm)	Aspartate α-keto

• Facts
  • Most sensitive enzyme for skeletal muscle disease
  • Inhibited by all anticoagulants except heparin (but ammonium heparin should not be used)

3. LD

• Sources
  • All tissues
• Methods
  • Wacker Method (forward)
  • Wrobleuski La Due (reverse)
  • Wrobleuski Cabaud
  • Berger Broida
• LD1 (anodic & heat stable)
• LD2 (heat stable & major isoenzyme in the sera of healthy persons)
• LD5 (cathodic & cold labile)
• Flipped ratio: LD1>LD2
• LD/HBD(LD1) ratio: 1.2-1.6;
• if 0.8-1.2 suspect for MI

4. Myoglobin

• Facts
  • Responsible for O2 supply of striated muscle

5. Troponin

• Facts
  • 3 subunits:
    • I (inhibitory)
    • T (tropomyosin-binding) 
    • C (calcium-binding)

Acute Pancreatitis Profile

1. Amylase

• Sources
  • Salivary glands
  • Pancreas

• Methods	Substrate
  Saccharogenic Iodometric/Amyloclastic Chromogenic Kinetic Method	Pancreatic AMS: diastase Salivary AMS: ptyalin

• Facts
  • MicroAMS: unbound (free)
  • MacroAMS: bound to IgG and IgA
  • Earliest pancreatic marker
  • Smallest enzyme in size
  • Salivary AMS: inhibited by wheat germ lectin

2. Lipase

• Sources
  • Pancreas

• Methods	Substrate
  Cherry-Crandall Sigma-Tietz Titration	Olive Oil/Triolein (pure form of TAG)

• Facts
  • End product: Fatty Acids
  • Most specific pancreatic marker

Prostatic Cancer Profile

1. ACP

• Sources
  • RBC
  • Prostate

• Methods	Substrate
  Chemical Inhibition Test RBC- ACP: inactivated by Cu++, unaffected by Tartrate P-ACP: unaffected by Cu++, inactivated by Tartrate	Organophosphates

• Facts
  • Very labile (add 5M acetate buffer/citrate tablet to preserve)

2. PSA

• Facts
  • Most useful for tumor marker for prostate cancer
  • RR: 0-4ng/mL