Tutorial:Basic Chemistry Topics

Possible Acetylation of Mycobacterium Tuberculosis by Tobramycin CoA Complex

Throughout this tutorial we will be targeting basic chemistry topics. The chemistry topics are based off a study conducted by a group of scientists. These topics are vital to the understanding of more advanced chemistry. There are interactive molecules incorporated into the text to help your understanding.

The study where this molecule was obtained is named "Aminoglycoside 2'-N-acetyltransferase from Mycobacterium tuberculosis-Complex with Coenzyme A and Tobramycin". The study focused on AAC (2’)- Ic also known as aminoglycoside 2’- N- acetyltransferase. The scientist’s in the study determined the crystal structure of AAC (2’)-Ic from Mycobacterium tuberculosis. The specific fold of AAC (2’)-Ic places in the GNAT or GCN5-related N-acetyltransferase superfamily. Although the physiological function of AAC(2’)-Ic in not certain, the crystal structure they determined allowed them to hypothesize. Through the crystal structure they determined that this enzyme might acetylate mycothiol. Mycothiol is key biosynthetic intermediate and the major reducing agent in mycobacterium. This enzyme is capable of acetylating aminoglycosides bearing a 2’ amino group, when this occurs the aminoglycoside antibiotic becomes inactive.

I know the summary above is complex and confusing. I don't expect you to understand it completely. I am sharing this study with you because I am going to use it to explain basic chemistry concepts. We are going to dissect this scientific artical and pull the basic concepts from it and go into greater detail.

Objectives

By the end of this tutorial you should be able to:

1. Understand and explain the importance of Tobramycin as an antibiotic

2. Describe and provide examples of covalent bonds, ionic bonds, and hydrogen bonds

Tobramycin

Tobramycin is an antibiotic part of the aminoglycoside family. Aminoglycosides produce antibacterial effects by inhibiting protein synthesis and compromising the cell wall structure. By inhibiting the protein synthesis of the bacteria it does not allow the bacteria to replicate. The cell wall is an important structure to bacteria, because it provides the structure and stability to the bacteria. By disrupting the cell wall we are removing the stability of the bacteria and ultimately casing bacteria death. Tobramycin targets a variety of bacteria particularly gram(-) species. Just like all drugs there are side effects associated with tobramycin. Some of the more common side effects are ototoxicity and nephrotoxicity. Ototoxic is hearing loss and nephrotoxic is causing kidney damage. The kidney damage is due to Tobramycin reabsorption through the renal tubules. This basically means that tobramycin may be toxic to the kidneys and the toxicity is caused by the contact-time in the renal tubules where the drug is located. Tobramycin trade name is Tobrex. A trade name is another name for tobramycin. It is a pregnancy category D. Pregnancy categories are assigned to all drugs. They are used to classify how likely the drug is to cause harm to the fetus. The pregnancy categories are A, B, C, D, and X. Pregnancy category A causes no harm to the fetus and pregnancy category X, which indefinitely causes harm to the fetus. Since Tobramycin is a pregnancy category D, this is not an optimal choice for a pregnant patient. Tobramycin can be given intravenously, intramuscularly, as an inhalation or ophthalmicly. Intravenously is an IV route of administration where the drug is administered directly to the vasculature or blood vessels. Intramuscular is a shot that penetrates your muscle. A common example of an intramuscular administration would be a flu shot. Inhalation is a route of administration where the lungs are the targets. An example of this would be an inhaler used in asthmatics. Ophthalmic administration is where the drug is administered to the eye; an example would be an eye drop.

Types of Bonds

There are 3 common types of bonds. A hydrogen bond, covalent bonds, or an ionic bond. The strongest bond is a covalent bond followed by the ionic bond, leaving the weakest bond to be the hydrogen bond. Covalent bonds, the strongest type of bond, they involves the sharing of electrons between two molecules. An example of a covalent bond is hydrochloric acid or HCl. The electrons are being shared between the chlorine atom (Cl) and the hydrogen atom (H). An ionic bond is an attraction between two molecules of opposite charge. The opposite charges I am referring to are a positive (+) and a negative charge (-). A positively charged atom is referred to as a cation, and a negatively charged atom is referred to as an anion. Hydrogen Bonds, the weakest of bonds, are attractive interactions (dipole-dipole) between an electronegative atom and hydrogen. Electronegative atoms are atoms that have high electron density. They are strong atoms that pull electrons towards then from weaker/low electron density atoms, such as hydrogen. When the electronegative atom pulls the electrons it leaves the other atom with a slight positive charge. The most common example of hydrogen bonding is water. The water molecule chemical formula is H2O. The highly electronegative oxygen pulls the hydrogen closer by attracting hydrogen’s electrons allowing the formation of a water droplet. The electronegative atoms allow for the droplet to be held together instead of spreading. The hydrogen bonds in this picture are displayed as yellow dashed lines. The hydrogen bonds in this molecule are important to the secondary structures providing the stability of the atoms orientation.

Secondary Structures

This molecule represents the . The alpha helices are represented with pink arrows and the beta strands are represented with yellow arrows. This molecule has approximately four alpha helices and two beta strands. The structure of the alpha and beta sheets in tuberculosis/CoA/ and Tobramycin structure represents the GNAT fold. The GNAT fold is almost completely catalyzed by CoA- dependent transfer of an acyl group to an exposed amino acid group. This interaction is the basis for the study. This shows how the acetylation of Mycothiol is occurring and why.

Active Site

The active site of a molecule can be described as a pocket where interaction between structures causes a desired effect. This is a good representation of the active site. The active site is where the substrate, in this case tobramycin, binds to CoA and the mycobacterium to cause an antibacterial effect. It the study described this is where the acetylation of the mycothiol should be occurring.

Ligand

The ligands displayed in the molecule to the right are and 3'-Phosphate-Adenosine-5'-Diphosphate. Coenzyme (CoA) is a coenzyme that synthesizes and oxidizes fatty acids. PAP's IUPAC name is [(2R,3S,4R,5R)-5-(6-aminopurin-9-yl)-4-hydroxy-3-phosphonooxyoxolan-2-yl]methylphosphono hydrogen phosphate. The ligands are held together by the hydrogen bonds created by alpha and beta sheets. The Protein in this molecule is represented as a dimer. A dimer is a chemical structure formed from two subunits. The dimer is constructed by connecting two subunits along their axis so that all four modules contribute to the structure.

This molecule displays the protein bound to the ligand CoA. The red molecules represent an anionic or negatively charged interaction. The dark blue molecules emphasize the cationic or positively charged interactions. The cationic and anionic interactions are contributed to arginine, aspartic acid, or glycine amino acids. The light blue molecules represent histidine, which is a basic amino acid. The difference in the charges displayed here contribute to the stability of the molecule. Since the charges are different it allows the molecules to be attracted to the opposite charge holding the molecule in a stable position.

Amino acids are the building blocks of proteins. There are 20 common amino acids. The contain and amine group (-NH2), a carboxylic acid group (-COOH) and a functional group specific to each amino acid. The functional group determines how the amino acid is classified. They are categorized as either, polar, non-polar, acidic or basic. There are 8 different amino acids present in the . CoA has a combination of 7 amino acids bound to it. The amino acids are two Arginine (basic amino acid), one Glycine (polar amino acid), and four Valine (non-polar amino acid). PAP has four amino acids bound to it, two Histidine and two tryptophan (non-polar amino acid). Tobramycin also has four amino acids bound to it, two aspartic acid (acidic amino acid)), Serine (polar amino acid) and tryptophan (non-polar amino acid)

CoA Amino Acids:

In this representation it displays the covalent bond between CoA and Arginine 124. Arginine is displayed as the pink molecule and CoA is displayed as the orange and red molecule. Arginine classified as a basic amino acid and is a nonessential alpha amino acid, meaning that can be synthesized by the human body.

Val96 and CoA are bound by a hydrogen bond, clearly displayed in this representation.

PAP Amino Acids:

This shows His54 bound to bound through a hydrogen bond.

The other Amino acid bound to PAP with a hydrogen bond is tcp

• References:

<Vetting, M. W., et al. "Aminoglycoside 2'-N-acetyltransferase from Mycobacterium tuberculosis-Complex with Coenzyme A and Tobramycin." RCSB Protien DataBase. N.p., 28 Aug.2002. Web. 13 July 2011. <http://www.rcsb.org/pdb/explore/explore.do?structureId=1M4D>.>