Enzymes

Formi

Enzymes

Hastens chemical reactions in organic matter

TRUE

TRUE/FALSE: The lower the enzyme concentration, the slower the reaction

FALSE

TRUE/FALSE: Higher concentration of substrate results in increased rate of reaction

Cofactors

Nonprotein entities that must bind to particular enzymes before a reaction occurs

NAD and NADP

Examples of coenzymes

Coenzymes

Essential to achieve absolute enzymatic activity

Activators

Inorganic ions which alter the spatial configuration of enzymes

Calcium, Magnesium, Zinc, Chloride, Potassium

Examples of activators

Metalloenzymes

Inorganic ions attached to a molecule

Catalase and Cytochrome Oxidase

Examples of metalloenzymes

Competitive Inhibitor

Physically binds to the active site of enzyme

Competitive Inhibitor

Inhibition is reversible when the substrate concentration is significantly higher than the inhibitor concentration

Add Excess Substrate

Competitive inhibition is counteracted by

Competitive Inhibitor

Has the ability to alter the apparent Michaelis-Menten constant

Dilution of Serum

Results to the reduction in the concentration of this inhibitor, thus increasing the rate of reaction

Non-Competitive Inhibitor

Look for areas other than the active site

TRUE

TRUE/FALSE: In non-competitive inhibition, the substrate and inhibitor may bind simultaneously

FALSE

TRUE/FALSE: In non-competitive inhibition, increasing the substrate concentration will reverse the inhibition

Uncompetitive Inhibitor

Binds to the enzyme-substrate complex

Increased Inhibition

Increasing the substrate in uncompetitive inhibition will

Isoenzymes

Enzymes that have the same catalytic reactions bu slightly different molecular structures \-- difference in the amino acid sequence

25C, 30C, 37C

Enzyme activities demonstrated at these temperature

37C

Optimum temperature for enzymatic activity

Increasing Temperature

Increase in molecule movement

40C to 50C

Temperature at which denaturation of enzymes is significant

60C to 65C

Temperature at which enzymes are inactivated

Temperature Coefficient

Every 10C increase in temperature, there will be a two-fold increase in enzyme activity

-20C

Ideal temperature for preservation of enzymes in a long period of time

2C to 8C

Ideal storage temperature for substrates and coenzymes

22C or Room Temperature

Ideal storage for LD (LD4 and LD5)

Hemolysis

Mostly increases enzyme concentration

Lactescence/ Milky Specimen

Decreases enzyme concenration

First Digit

Enzyme Classification

Second and Third Digits

Represents the subclass to which the enzyme is assigned

Fourth and Final Digit

Serial Number that is specific to each enzyme

EC 3.1.3.2

EC: Acid Phosphatase (ACP)

EC 3.1.3.1

EC: Alkaline Phosphatase (ALP)

EC 3.2.1.1

EC: Amylase

EC 2.6.1.2

EC: Alanine Aminotransferase (ALT)

EC 2.6.1.1

EC: Aspartate Aminotransferase (AST)

EC 4.1.2.13

EC: Aldolase

EC 3.4.15.1

EC: Angiotensin Converting Enzyme

EC 2.7.3.2

EC: Creatine Kinase

EC 3.1.1.7

EC: True/Acetyl Cholinesterase

EC 3.1.1.8

EC: Pseudocholinesterase

EC 2.3.2.2

EC: Gamma Glutamyl Transferase (GGT)

EC 1.1.1.49

EC: Glucose-6-Phosphate Dehydrogenase (G-6-PD)

EC 3.1.1.3

EC: Lipase

EC 1.1.1.27

EC: Lactate Dehydrogenase (LDH)

EC 3.1.3.5

EC: 5' Nucleotidase (5'N)

Oxidoreductase

Catalyze the removal or addition of electrons (Red-Ox Reactions)

CO, LDH, MDH, ICD,G-6-PD

Examples of Oxidoreductases

Transferases

Catalyze the transfer of a chemical group other than hydrogen

CK, AST, ALT, OCT, GGT

Examples of Transferases

Hydrolases

Catalyze the hydrolysis or splitting of a bond by the addition of water

Esterases (ACP, ALP, CHS, LPS)
Peptidases (Trypsin, Pepsin, LAP)
Glycosidases (AMS, Galactosidase)

Examples of Hydrolases

Lyases

Catalyze the removal of groups from substrates without hydrolysis

Glutamate, Pyruvate, and Tryptophan Decarboxylase, and Aldolase

Examples of Lyases

Isomerases

Catalyze the intramolecular arrangement of the substrate compound

Glucose Phosphate Isomerase and Ribose Phosphate Isomerases

Examples of Isomerases

Ligases

Catalyze the joining of two substrate molecules

Synthase

Example of Ligases

Active Site

Water-free cavity

where the substrate interacts

Allosteric Site

Cavity other than the active site

Prosthetic Group

A coenzyme that is tightly bound to the enzyme

Holoenzyme

Apoenzyme + Prosthetic Group

Proenzyme/ Zymogen

Inactive form of an enzyme

Emil Fischer's/ Lock and Key Theory

Based on the premise that the shape of the key (substrate) must into the lock (enzyme)

Kochland's/ Induced Fit Theory

Based on substrate binding to the active site of the enzyme

Absolute Specificity

An enzyme combines only with one substrate and catalyzes only one reaction

Group Specificity

Enzyme combines with all the substrates in a chemical group

Bond Specificity

Enzymes reacting with specific chemical bonds

Zero-Order Reaction

The reaction rate depends only on enzyme concentration

First-Order Reaction

The reaction rate is directly proportional to substrate concentration

Fixed-Time/ Endpoint Assay

Reactants are combined

the reaction proceeds for a designated time

the reaction is stopped and measurement is made

Continuous Monitoring/ Kinetic Assay

Multiple measurements of change in absorbance are made during the reaction
Preferred over fixed-time/endpoint assay

1 micromole of substrate/ minute

International Unit (IU or U)

1 mole of substrate/ second

Katal Unit

Alkaline Orthophosphoric Monoester Phosphohydrolase

AKA Alkaline Phosphatase

Alkaline Phosphatase

Functions to liberate inorganic phosphate from an organic phosphate ester with the production of an alcohol

Liver, Bone, Placenta, Intestine

Major Tissue Sources of Alkaline Phosphatase

30-90 U/L

Reference Value for ALP

Liver ALP, Bone ALP, Placental ALP, Intestinal ALP

Major Isoenzymes of Alkaline Phosphatase

Liver and Bone

Normal sera ALP is derived from

Bone ALP

ALP Isoenzyme increased in children (periods of growth) and adults \>65 years old

Group A and AB

Placental ALP has lower levels in pregnant patients with this blood group/s

16 - 20 weeks

In normal pregnancy, increased ALP is detected at

Group B and O

Higher levels of ALP due to differences in intestinal ALP levels

Blood Group (Secretor and H Substance)

Intestinal ALP Isoenzyme in serum depends on

Increase serum ALP and Creatinine

Period of Growth and Muscle Development

Liver ALP Fraction

Major contributor to increased Total ALP
Due to obstructive jaundice

Paget's Disease (Osteitis Deformans)

Highest elevation of Alkaline Phosphatase

B1x Isoform

Detected in serum of dialysis patients

B1x Isoform

Used to study Low Bone Mineral Disease (BMD) in patients with Chronic Kidney Disease

Regan ALP

Found in the lung, breast, ovarian, and gynecological cancers

65C for 30 Minutes

Heat stability of Regan ALP

Regan ALP

Co-migrator of Bone ALP
Inhibited by phenylalanine

Nagao ALP

Found in adenocarcinoma of the pancreas and bile duct, pleural cancer

Nagao ALP

Variant of Regan ALP
Inhibited by L-leucine and phenylalanine

Liver and Bone ALP

Most anodal ALP isoenzyme