# Structural Biochemistry/Enzyme/Cofactors

## General information

Cofactors, mostly metal ions or coenzymes, are inorganic and organic chemicals that assist enzymes during the catalysis of reactions. Coenzymes are non-protein organic molecules that are mostly derivatives of vitamins soluble in water by phosphorylation; they bind apoenzyme to proteins to produce an active holoenzyme. Apoenzymes are enzymes that lack their necessary cofactor(s) for proper functioning; the binding of the enzyme to a coenzyme forms a holoenzyme. Holoenzymes are the active forms of apoenzymes.

holoenzyme
cofactor

Cofactors can be metals or coenzymes, and their primary function is to assist in enzyme activity. They are able to assist in performing certain, necessary, reactions the enzyme cannot perform alone. They are divided into coenzymes and prosthetic groups. A holoenzyme refers to a catalytically active enzyme that consists of both apoenzyme (enzyme without its cofactor(s)) and cofactor. There are two groups of cofactors: metals and small organic molecules called coenzymes. Coenzymes are small organic molecules usually obtained from vitamins. Prosthetic groups refer to tightly bound coenzymes, while cosubstrates refer to loosely bound coenzymes that are released in the same way as substrates and products. Loosely bound coenzymes differ from substrates in that the same coenzymes may be used by different enzymes in order to bring about proper enzyme activity.

General formula

## Metal cofactors

Metal ions are common enzyme cofactors. Some enzymes, referred to as metalloenzymes, cannot function without a bound metal ion in the active site. In daily nutrition, this kind of cofactor plays a role as the essential trace elements such as: iron (Fe3+), manganese (Mn2+), cobalt (Co2+), copper (Cu2+), zinc (Zn2+), selenium (Se2+), and molybdenum (Mo5+). For example, Mg2+ is used in glycolysis. In the first step of converting glucose to glucose 6-phosphate, before ATP is used to give ADP and one phosphate group, ATP is bound to Mg2+ which stabilizes the other two phosphate groups so it is easier to release only one phosphate group. In some bacteria such as genus Azotobacter and Pyrococcus furiosus, metal cofactors are also discovered to play an important role. An example of cofactors in action is the zinc-mediated function of carbonic anhydrase or the magnesium-mediated function of restriction endonuclease.

## Coenzyme

A coenzyme is a small, organic, non-protein molecule that carries chemical groups between enzymes. It is the cofactor for the enzyme and does not form a permanent part in the enzyme's structure. Sometimes, they are called cosubstrates and are considered substrates that are loosely bound to the enzyme. In metabolism, coenzymes play a role in group-transfer reactions, such as ATP and coenzyme A, and oxidation-reduction reactions, such as NAD+ and coenzyme Q10. Coenzymes are frequently consumed and recycled. Chemical groups are added and detached continuously by an enzyme. ATP synthase enzyme phosphorylates and converts the ADP to ATP, while Kinase dephosphorylates the ATP back to ADP at continuous rates as well. Coenzyme molecules are mostly derived from vitamins. They are also commonly made from nucleotides such as adenosine triphosphate and coenzyme A.

Through further research in coenzyme activity and its binding effect on the enzyme, more can be revealed about how the enzyme changes conformationally and functionally. An example is of the MAPEG group of integral membrane enzymes. These enzymes are crucial in the catalytic transformation of lipophilic substrates, which are involved in arachidonic acid derived messengers production and xenobiotic detoxification. Through use of a bound detergent to mimic a MAPEG enzyme's cofactor, glutathione, a new active site specific for lipophilic substrate is revealed; thus, further studies can reveal how these substrates bind to this second form of the enzyme [1].

Vitamin C is an important coenzyme
Vitamin A

## Important Coenzymes

nicotinamide adenine dinucleotide is a coenzyme derived from vitamin B3. In NAD+ the functional group of the molecule is only the nicotinamide part. NAD+ is capable of carrying and transferring electrons and functions as oxidizing agent in redox reactions. It also works as a substrate for DNA ligases in posttranslational modification, where the reaction removes acetyl groups from proteins. Furthermore, in glycolysis and the citric acid cycle, NAD+ oxidizes glucose and releases energy, which is then transferred to NAD+ by reduction to NADH. NADH later on unloads the extra electron through oxidative phosphorylation to generate ATP, which is the energy source humans use every day. In addition to catabolic reactions, NADH is also involved in anabolic reactions such as gluconeogenesis, and it also aids in the production of neurotransmitters in the brain.

flavin adenine dinucleotide is a prosthetic group that, like NADH, functions as a reducing agent in cellular respiration and donates electrons to the electron transport chain.

### Quinone

compounds that have fully conjugated aromatic rings to which two oxygen atoms are bounded as carbonyl groups (i.e. diketones). Quinone’s structure gives them the ability to form substances with colors. They exist as pigments in bacteria, fungi, and certain plants, and give them their characteristic colors. In addition, they are used to manufacture different color dyes for industrial purposes. In biological systems, they serve as electron acceptors (oxidizing agents) in electron transport chains such as those in photosynthesis and aerobic respiration. Many natural or synthetic quinines show biological or pharmacological activities, and some event show antitumoral activities.

### CoA

coenzyme A, synthesized from pantothenic acid ATP, functions as acyl group carriers to transport functional groups such as acetyl (acetyl-CoA) or thioesters in metabolic reactions like fatty acid oxidation (synthesis of fatty acids) and citric acid cycle (cellular respiration). It also transfers fatty acids from cytoplasm to mitochondria. In addition to its transporter role in metabolism, CoA is also an important molecule in itself. For instance, CoA is an important precursor to HMG-CoA, an important enzyme in the metabolic synthesis of cholesterol and ketones. Furthermore, it contributes the acetyl group to the structure of acetylcholine, which is an important neurotransmitter responsible for inducing muscle contraction.

## Common Coenzymes

### Vitamin A

Vitamin A is subdivided into two molecules, Vitamin A1 (retinol) and Vitamin A2 (dehydroretinol). Retinol is the most active and common form. Vitamin A has a large conjugated chain which serves as the reactive site of the molecule. Unlike most cofactors, Vitamin A undergoes a sequence of chemical changes (oxidations, reductions, and isomerizations) before returning to its original form. The ability for Vitamin A's electrons to travel from ${\displaystyle \pi {\text{ to }}\pi ^{*}}$ orbital makes it a good candidate molecule for trapping light energy. Consequently, Vitamin A is responsible for transferring light energy to a chemical nerve impulse in the eyeball. Vitamin A is also used for growing healthy new cells such as skin, bones, and hair. It maintains the lining of the urinary tract, intestinal tract, and respiratory system. Additionally, Vitamin A is required for the reproductive functions such as the growth and development of sperm and ovaries.

### Vitamin C

Also known as ascorbic acid, Vitamin C is quite abundant in most plants and animals excluding primates, guinea pigs, bats, and some birds. Despite human's inability to synthesis absorbic acid, it is an essential in many biosynthetic pathways such as synthesizing collagen. Deficiency leads to a disease called Scurvy. Vitamin C helps regulate the immune system and relieve pain caused by tired muscles. It also is needed in the manufacture of collagen and norepinephrine. Vitamin C is also an antioxidant which can enhance the immune system by stimulating white blood cells in the body. Vitamin C also helps to benefit the skin, teeth, and bones.

### Vitamin B1

Also named Thiamine or Thiamine diphosphate (TPP), Vitamin B1 is a cofactor for oxidative decarboxylation both in the Kreb's Cycle and in converting pyruvate to acetyl-CoA (an important molecule used in the citric acid cycle of metabolism). It is widely available in the human diet and particularly potent in wheat germ and yeast. It's functionality results from a thiazole ring which stabilizes charge and electron transfer through resonance.

### Vitamin B2

Vitamin B2 is known as riboflavin. Vitamin B2 is the precursor of Flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN) which are coenzymes used to oxidized substrates. FAD contains riboflavin and adenine. FMN contains riboflavin that is why it is called mononucleotide.

### Vitamin B3

Vitamin B3 is Niacin or nicotinic acid with the formula C5H4NCO2H. Vitamin B3 is a precursor to NADH, NAD+, NADP+ and NADPH which are coenzymes found in all living cells. NAD+ and NADP+ are oxidizing agents. NADH and NADPH are reducing agents.

### Vitamin B6

Vitamin B6 is precursor to coenzyme pyridoxal phosphate (PLP) which is required in certain transformation of amino acids including transamination, deamination, and decarboxylation.

### Vitamin B12

Vitamin B12 is the name for a class of related compounds that have this vitamin activity. These compounds contain the rare element cobalt. Humans can not synthesize B12 and must obtain it from diet.

### Vitamin H

Also named Biotin, Vitamin H is a carboxyl carrier; it binds CO2 and carries it until the CO2 is donated in carboxylase reactions. It is water soluble and important in the metabolism of fatty acids and the amino acid Leucine. Deficiency leads to dermatitis and hair loss, thus making it a popular ingredient in cosmetics.

### Vitamin K

Vitamin K is needed for the process of clotting of blood and Ca2+ binding. Vitamin K can be synthesized by bacteria in the intestines. Vitamin K is needed for catalyzing the carboxylation of the γ-carbon of the glutamate side chain in proteins.

## Non-enzymatic cofactors

Cofactor is also used widely in the biological field to refer to molecules that either activate, inhibit or are required for the protein to function. For example, ligands such as hormones that bind to and activate receptor proteins are termed cofactors or coactivators, while molecules that inhibit receptor proteins are termed corepressors.

The coactivator can enhance transcription initiation by stabilizing the formation of the RNA polymerase holoenzyme enabling faster clearance of the promoter.

The corepressor can repress transcriptional initiation by recruiting histone deacetylases which catalyze the removal of acetyl groups from lysine residues. This increases the positive charge on histones which strengthens in the interaction between the histones and DNA, making the latter less accessible to transcription.

# References

1. Dewick, Paul. Medicinal Natural Products: A Biosynthetic Approach. 3rd ed. West Sussex, Britain: Wiley, 2009. 32-34. Print.
2. von Heijne, G and Rees,D (2008). Current Opinion in Structural Biology. Elsevier Ltd.
1. von Heijne G, Douglas R: Membranes:reading between the lines. Current Opinion in Structural Biology 2008, 18:403-405