Cofactors

• Cofactors are organic or inorganic molecules that are required for the activity of a certain conjugated enzymes

• Apoenzyme (inactive): enzyme without cofactor

• Holoenzyme (active): enzyme with cofactor

• Inorganic cofactors: essential ions

• Organic cofactors: coenzymes

I. Essential Ion Cofactors

• Metal activated enzymes – require or are stimulated by addition of metal ions - usually monovalent cations (e.g. K+, Na+) and divalent cations (e.g. Mg2+, Zn2+)

• Metal ions of metalloenzymes – cations that are tightly bound to enzyme and participate directly in catalysis – usually transition metals (e.g. Fe, Zn, Cu, Co)

Metalloenzymes (with bound metal ions at active sites)

Iron sulfur clusters in enzymes involved in oxidation-reduction - Fe complexed with S - Accept or donate electrons

Carbonic anhydrase - A Zn2+ metalloenzyme - CO2 + H2O → bicarbonate

CO2 H2O

II. Coenzymes

Cosubstrates- - altered in a reaction and dissociated from active site - regenerated to original structure in a subsequent reaction - transfer chemical groups among different enzyme reaction. Prosthetic groups- - remains bound to enzyme - must return to original form Both cosubstrates and prosthetic groups supply reactive groups not

present on the active site amino acid side chains

Vitamin derived coenzymes

• Must be obtained from diet • Synthesized by microorganisms and plants • Vitamin deficiencies lead to disease state • Most vitamins must be enzymatically transformed to function as a

coenzyme

What about vitamin B4 and B8?

1. ATP and other nucleotide coenzymes - Supply chemical group to other substrates

A. ATP:

Phosphoryl groups

Nucleotidyl group (AMP)

ATP + substrate

ATP + substrate

ATP + substrate

Substrate–P + ADP

Substrate–PP + AMP

Substrate–AMP + PPi

B. S-Adenosylmethionine (SAM):

Methyl group

Methionine + ATP SAM

SAM

SAM is a donor of methyl group in biosynthetic reaction

Example:

C. Nucleotide sugar phosphates - Involved in carbohydrate metabolism - UDP-glucose: carrier of glucose for transfer to other substrate

Formation of UDP-glucose:

Glucosyl group

2. NAD+ and NADP+

- Nicotinamide coenzymes - Nicotinamide adenine dinucleotide (phosphate) - Precursor – nicotinic acid (Niacin, B3) - Tryptophan can be degraded to niacin - Oxidation-reduction reactions

3. FAD and FMN

Riboflavin

- Flavin coenzymes - Flavin adenine dinucleotide (FAD); Flavin mononucleotide (FMN) - Precursor – riboflavin (B2) - Prosthetic group - Oxidation-reduction reactions

FAD

= H in FMN

4. Coenzyme A and acyl carrier protein (ACP) - Transfer of simple carboxylic acids and fatty acids - Free thiol (-SH) group for acyl group attachment

Co-enzyme A

ACP

Vitamin B5

5. Thiamine Diphosphate (TDP or TPP) - Made from thiamine (B1) - Prosthetic group for enzymes in decarboxylation/carboxylation

- first Vitamin discovered (Vital amine = Vitamin)

TDP in decarboxylation of pyruvate to acetylaldehyde:

6. Pyridoxal 5’-phosphate (PLP) - Made from Vitamin B6 - Prosthetic group for enzymes catalyzing reactions involving amino acids

Vitamins of the B6 family

PLP in transamination reaction

1 1

7. Biotin - Vitmain B3 - Carboxyl group transfer reaction - ATP-dependent carboxylation - Prosthetic group - Bound to lysine residue in enzyme

Biotin in ATP-dependent carboxylation of pyruvate

8. Tetrahydrofolate (THF)

- Folate is converted to THF by the addition of 4 hydrogens to the pterin ring. - Important in transfer of one-carbon units (N10 and N5 are involved) - THF Has a poly-glutamate tail formed by gamma-carboxyl and alpha amino groups (unusual peptide bond).

β α γ β α γ

One-carbon derivatives of THF:

9. Cobalamin (B12) coenzymes - B12 is the largest B vitamin - Corrin ring with cobalt cation - Coenzymes: methylcobalamin and adenosylcobalamin

- Participate in reactions involving intramolecular rearrangements, methyl group transfer, etc.

Cobalamin coenzymes

1. Intramolecular rearrangement:

2. Methyl group transfer:

5-methylTHF THF

Cobalamin Methylcobalamin

Methionine Homocysteine