Haem/kinetics/mitochondria (Question 1-14)

Front 1

What makes up Haemoglobin (Basics)?

Back 1

Protoporphyrin IX, Iron (Fe^2+). Quarternary structure, 4 polypeptide chains, 2 dimers.
Polypep'chains are held together by NON-COVALENT interactions, Dimers by HYDROPHOBIC (Ionic/H-bonds).

Front 2

Hb-A

Back 2

2alpha - 2beta;
Main form of Haemoglobin found in a healthy adult.

Front 3

Hb-F

Back 3

2alplha - 2gamma;
Found in the human fetus;
60% of total haem in the human fetus in the last few months of fetal life;
Hb-A synthesis starts at the 8th month;
Higher O2 affinity than Hb-A.

Front 4

Hb-A2

Back 4

2alpha - 2delta;
2% of total haemoglobin found in a healthy adult.

Front 5

Hb-A1c

Back 5

Slightly glycosylated haemoglobin;
A non-enzymatic reaction occurs between glucose and the N-end of the beta chain;
More frequent in patients with Diabetes Mellitus.

Front 6

Methemoglobin

Back 6

Oxidation of the Haem;
Fe^2+ goes to Fe^3+ = cannot bind to oxygen;
Drugs can cause..
A lack of NADH-Cytochrome B5 Reductase can stop converting Methemoglobin back into Haem - meaning trouble for the individual = Tissue Hypoxia.. due to lack of oxygen supply.
CHOCOLATE CYANOSIS - Treated with METHYLENE BULE.

Front 7

6 Enzyme Classes

Back 7

1.) Oxidoreductases - One substance is oxidized, the other is reduced;
2.) Transferases - Transfer of the functional group, from one substance to another;
3.) Hydrolases - C-O/C-N/C-S.. either one of these bonds being broken by the addition of H2O;
4.) Lyases - C-C/C-O/C-N/C-S.. either one of these bonds bying broken by other means than OXIDATION/HYDROLYSIS;
5.) Isomerases - Rearrangement of existing molecules;
6.) Ligases - C-C/C-O/C-N/C-S.. either one of these bonds in reactions, in conjunction with the cleavage of ATP or other high energy compound..

Front 8

What is an Active Site?

Back 8

The active site is usually found in a 3-D groove/pocket of an enzyme. It is lined with amino acid side chains (or nucleotides in RNA enzymes). These side chains are involved in recognition of the substrate. After an active site has been involved in a reaction, it can be used again.

Front 9

Isoenzymes?

Back 9

Differ in amino acid sequence but are able to catalyse the same chemical reaction;
CK-MM = Muscle; 94% (Total activity in blood plasma)
CK-MB = Cardiac; >6%
CK-BB = Brain. Trace

Front 10

V = K[S] = K[S]^1

Back 10

Reaction of First Order.
As the concentration of the substrate DECREASES, the concentration of the product INCREASES.

Front 11

V = K[S]0 = K x 1 = K

Back 11

Reaction of Zero Order.
Reaction rate DOES NOT depend on the substrate concentration.

Front 12

Michaelis-Menten Equation

Back 12

V0 = Vmax x ( [S]/ [S] + Km )

Front 13

[S] < Km =
[S] > Km =
[S] = Km =

Back 13

1.) LOW Substrate concentration.. reaction = 1st Order;
2.) HIGH Substrate concentration.. reaction = 0 Order;
3.) Vmax x ( [S]/ [S] + [S] ) = Vmax x ( [S]/2[S] ) = Vmax/2.

Front 14

Enzyme Activity?

Back 14

..measure of the ability of an enzyme to catalyze a specific reaction.

Front 15

What are the units for the 'Catalytic Activity of Enzymes'?

Back 15

Katal;
1 Katal = mol/s... can also be expressed in IU (International Unit);
1IU = 16.6nkat
1Ukat = 60IU

Front 16

Catalytic Concentration of Enzymes..

Back 16

Activity is related to the volume of the body fluid - Blood Serum;
mkat/l & ukat/l - these are typical units for it.

Front 17

Kinetic method of measuring 'Catalytic Concentration'..

Back 17

Either Substate/Product can be measured;
Continually measured;
Needs a KINETIC Curve;
Initial velocity is determined;
Exact determination (of value).

Front 18

Constant-Time method of measuring 'Catalytic Concentration'..

Back 18

Just Product can be measured;
After some time, reaction stops by enzyme inactivation - then measure;
Does not need a KINETIC Curve;
Average velocity is determined;
The value determined is not an exact value.

Front 19

Factors affecting the catalytic activity of enzymes..

Back 19

Temperature;
Activators/Inhibators;
Competitive inhibitors/Non-Competitive inhibitors..
(Comp'Inhib - INCREASES Km without any change in Vmax; Non-Comp'Inhib - DECREASES Vmax without any change in Km);
pH;
Substrate Concentration;
Enzyme Concentration.

Front 20

Metaloenzymes..

Back 20

These enzymes contain functioning metal cations - prosthetic groups - that are directly incolved in the catalyzed reaction.. Some enzymes are inactive without 'their' metal ion being present.

Front 21

Metaloenzyme..
Molybdenum (Mo)

Back 21

Xantine Oxidase (Xantine -- Uric Acid)
Sulfite Oxidase (Sulfite HSO3 -- Sulfate SO4^2-)

Front 22

Metaloenzyme..
Zinc (Zn)

Back 22

Alcohol Dehydrogenases (Ethanol -- Acetaldehyde);
Carbonic Anhydrase (H2O + CO2 <> H2CO3)'
Carboxypeptidases (Cleavage of Polypeptides from the C-Terminal).

Front 23

Metaloenzyme..
Copper (Cu)

Back 23

Ceruloplasmin (Ferrooxidase) - (Fe2+ -- Fe3+);
Cytochrome-C-Oxidase.

Front 24

Where is NAD+ needed?
(Pyridine Nucleotide-dependent dehydrogenases)

Back 24

CAC - Isocitrate Dehydrogenase, 2-Oxoglut Dehydrogenase, Malate Dehydrogenase;
Glycolysis - Glyceraldehyde 3P Dehydrogenase, Lactate Dehydrogenase;
Oxidation of Ethanol - Alcohol Dehydrogenase, Acetaldehyde Dehydrogenase..
Basically remember that Dehydrogenases require NAD+ and give off NADH + H^+ :).

Front 25

Where is FAD needed?

Back 25

Mainly required for the Dehydrogenation of Succinate to Fumarate.. found on the Respiratory Chain (Coenzyme 2, found on the Inner Mitochondrial Membrane) and also part of the CAC (Stage 8).

Front 26

What are cytochromes?

Back 26

They are Haem proteins that are one-electron carriers, due to the reversible oxidation of the Iron atom. Each cytochrome has a Haem group - made of Porphyrin Ring and an Iron atom.
The Iron atom is reversibly converted from Fe^3+ to Fe^2+ - part of its function as a reversible carrier of electrons.

Front 27

Cytochrome C

Back 27

Fe atom is attached via Coordination to N-Atom of His and S-Atom of Met.
The haem is deep within the tertiary structure of the protein, meaning it is unable to bind to O2 or CO.
MOVES ON THE OUTER SIDE OF THE INNER-MITO-MEMB.

Front 28

Cytochrome aa3

Back 28

Fe atom is attached via Coordination to two His residues, one is a hydrophobic isoprenoid chain, other is oxidized to formyl.
Haem a - acceptor of an electron from copper centre. CO inhibits, as do CN/HS and N3 (all minus) ions.

Front 29

Mitochondria

Back 29

Membrane enclosed organelle, double membrane, cristae, matrix. 0.5-1 micrometer in diameter. Supply most of the cells supply of ATP. Involved in Cell Signaling, Cell Differentiaton, Cell Death and partly in-charge of the cell cycle and cell growth.
Outer Membrane - Fairly permeable to small molecules/ions - VDAC are present (voltage dependent anion channel).
Inner Membrane - Basically impermeable to virtually all ions and polar molecules - loads of specific transporters are present (PYRUVATE, MALATE, CITRATE, ATP..).
Involved in:
HEAT PRODUCTION, STORAGE OF CALCIUM IONS, APOPTOSIS,
KREBS, FA DEGRADATION, GLYCOLYSIS, 1st/Last PARTS OF HAEM SYNTHESIS.