01 Building Blocks of tissues and cells

1 Building Blocks of tissues and cells

Before we follow the digestion and metabolism of nutrients, we need to discuss what building blocks are required to make a cell. Please note that our food is derived from plant cells or animals cells and therefore has the very same building blocks that are used to make our own cells.

Learning goals

Understand what is required to make a cell.
Appreciate the chemical structure of biomolecules.
Titration of amino acids.
Get a first impression of the purposes of biomolecules.

Key concepts

Dynamics of a living cell
Physico-chemical properties of amino acids
Physico-chemical properties of lipids
Building blocks of a cell

Building blocks of tissues and cells

Nutrition is needed to provide energy, but also to provide building blocks that make up tissues and cells. It is an instructive exercise to consider what compounds are required to build an organism. To make a new cell we need (Table 1):

Building block	Structure
Amino acids	Protein
Nucleotides	DNA, RNA
Lipids	Membranes, Storage
Carbohydrates	Storage, protein and lipid modification, nucleotides
Vitamins	Enzyme cofactors, antioxidants
Minerals, Metals	Enzyme cofactors, modification of molecules and proteins, cytosolic ions, oxygen transport, modification of molecules and proteins

Watch this video to get an impression of how cells work and how dynamic they are, when looked at a high magnification. You will not understand all the details at this point, but the animation shows an immune cell rolling along the wall of a blood vessel. The observer then moves to the membrane and looks into the cell and its cytoskeleton. Next you will look at the synthesis of protein using mRNA as a template. Then watch how some proteins are inserted into the membrane of the endoplasmic reticulum. The membranes with proteins bud off and move to the plasma membrane where they fuse and release contents.

We have ample reason to believe that every biological phenomenon, however complex, is ultimately based on chemical and physical interactions among molecules. Harold, Franklin M.. The Way of the Cell (p. 4). Oxford University Press. Kindle Edition.

In figure 1 and the video the cell looks quite empty. Is this real. Figure 2 shows an illustration of the crowded environment in a living cell.

Protein is quantitatively the largest component of any cell. It includes all enzymes to carry out metabolism, signalling proteins, defence proteins, structural proteins and storage proteins. All proteins are made up of 20 different amino acids (Fig. 3)

As the name suggests, amino acids have an α-amino-group and an α-carboxyl-group. The α-carbon atom to which these two functional groups are attached has two further bonds available. One is bound to hydrogen, the other can be a hydrogen (glycine) or extends into a larger sidechain, referred to as R- if not specified. The sidechain gives the amino acid distinct biophysical properties that allow amino acids to undergo specific interactions with other molecules or amino acids, such as hydrogen bonds, hydrophobic interactions, ionic bonds and disulfide bridges. Typically, amino acids are classified as neutral non-polar, neutral polar, aromatic, anionic (acidic) and cationic (basic). Proline is unusual among amino acids, because its amino-group is part of a pyrrolidine structure. This has significant structural consequences for the flexibility of larger structures formed by proline-containing peptides. Amino acids occur in two mirror-image forms, designated as L- or D- amino acids. For protein synthesis only L-amino acids are used. D-amino acids occur in nature and can be generated and/or metabolised through enzymatic conversion (from D- to L- and L- to D-) by amino acid racemases. D-serine, for example, is an important neuromodulator.

An important feature of amino acids is the dissociation of protons from suitable functional groups. Table 2 lists functional groups that occur in amino acids. The pKa values indicates at what pH the corresponding group is 50% protonated/50% deprotonated and therefore shows whether a functional group can act as an acid or base. The third column shows which amino acids have the corresponding functional group.

Table 2: Acid-base properties of amino acids

Functional group	pKa (approximate)	Amino acid
α-carboxyl-group	3.1	all
Terminal carboxyl-group	4.1	aspartate, glutamate
Imidazole	6.0	histidine
Sulfhydryl	8.3	cysteine
α-amino group	9.0	all
Phenylhydroxy-group	10.9	tyrosine
Terminal amino-group	10.5	lysine
Guanidinium	12.5	arginine

Watch the following video to see the titration of the amino acid glycine. In Figure 4 you can then look at the more complex acid/base properties of histidine, which has 3 ionizable groups.

To illustrate the change of amino acid protonation with pH, histidine is shown as an example (Fig. 4). When dissolved in 0.1M HCl (pH=1) all ionisable groups are protonated. Hence, the carboxyl-group will be –COOH, the amino-group will be –NH₃⁺ and one of the nitrogens in the imidazole ring will be protonated as well =NH⁺-. Due to the delocalisation of the positive charge in the imidazole ring, acidification does not protonate both nitrogens. To this solution 0.1M NaOH is added dropwise and the pH followed. At pH 5.0, the most acidic group (the α-carboxyl-group) will be fully dissociated to –COO^–. Further titration will cause dissociation of the proton associated with the imidazole ring (=N-), which is largely accomplished at pH 7.5. Titration to pH 10, will see dissociation of the proton from the α-amino group (-NH₂). The amino acid acts as a pH buffer at its pKa values. Let us use the pKa value of the α-carboxyl-group as an example. At pH 3.1, 50% of the α-carboxyl-group will be protonated (-COOH) and 50% will be deprotonated (-COO^–). Addition of protons will cause some of the -COO^– to become -COOH, thereby absorbing the protons that would make the solution more acidic. Vice versa adding OH^– will cause some -COOH to lose a proton, which neutralises the OH- to form water. Once this process is exhausted the buffering power ceases.

Notably, the listed pK (Table 2) values apply to free amino acids in aqueous solution. At first glance only histidine appears to have a side-chain the charge of which can change in the physiological range of pH 6-8. However, the pK-value of amino acid side-chains can change dramatically depending on the environment. For instance, a more hydrophobic environment favours functional groups to be uncharged. Thus, a carboxyl-group is forced to remain protonated (-COOH). As an example, the side-chain carboxyl-group of glutamate, which in aqueous solution has a pK = 4.1, is often found protonated in the active site of enzymes, where it is surrounded by other amino acid side-chains and water. As a result, certain glutamate residues can act as an acid or as a base in biochemical reactions. Amino acid side-chains are often classified as polar when they have the ability to form hydrogen bonds or charge pairs, while being classified non-polar when they cannot form these interactions (aliphatic side chains). Non-polar (hydrophobic) amino acids are generally found in the core of proteins where they can interact with other hydrophobic side-chains instead of being exposed to polar water molecules. For the same reason non-polar amino acids are also enriched in transmembrane helices. The polar amino acid cysteine is special because the sulhydryl-groups (-SH) of two adjacent cysteine residues can oxidise to form a covalent disulfide bridge (-S-S-) (golden in Fig. 5). This bond is much stronger than any of the weak interactions formed by other amino acids. Disulfide bridges can only form in oxidising environments outside the cell, such as the blood, where you would find the hormone insulin (Fig. 5). We will revisit the properties of amino acids in more detail when looking at protein structure.

Test your knowledge
Q: What functional groups are common to all amino acids.

Imidazole ring
alpha-amino group
delta-carboxyl group
Guanidinium group
alpha-carboxyl group

Show answer

2 and 5, alpha-amino group and alpha-carboxyl group.

Nucleotides

Nucleotides are the building blocks of RNA and DNA molecules. However, nucleotides also serve important additional role as metabolites. The RNA building block ATP (Adenosinetriphosphate) is the major mobile energy carrier in all organisms. GTP (Guanosinetriphosphate), while used less universal, is used as an energy carrier in protein biosynthesis and in signal transduction processes. CTP (Cytidinetriphosphate) and UTP (Uridinetriphosphate) are used as energy carriers for carbohydrate conjugation. Nucleotides are a compound building block, made up of a carbohydrate (ribose or deoxyribose), a nucleobase (adenine (A), thymine (T), guanine (G), cytosine (C), and uracil (U)) and phosphate groups. In RNA ribonucleottides are used while in DNA deoxyribonucleotides are used.

The naming convention is adenine, guanine, cytosine, thymine and uracil for the nucleobases. When attached to ribose they are called nucleosides and named adenosine, thymidine, guanosine, cytidine and uridine. They are called nucleotides when they contain phosphate groups in addition, which are attached to the C5 of ribose. To avoid confusion with the carbon atoms in the nucleobase the numbering is used with a prime e.g. 5’ or 3’.

This can be used to indicate the direction of DNA polymerization. In DNA, 2-deoxyribose is used in nucleotides and A,T,G,C are used as nucleobases. In RNA ribose is used and A,U,G,C are the bases. The chemical nature of the nucleobases allows the formation of parallel hydrogen bonds with 2 bonds occurring between A and T and 3 bonds occurring between G and C. It is worth noting that nucleobases are planar, but there is a distinct angle in the bond to ribose, which allows adoption of the well-known helical structure by complementary strands of oligonucleotides. All cells can synthesize nucleotides de novo but are also able to salvage nucleotides from the circulation.

Lipids

Membranes enclose all cells and also their sub-compartments, such as lysosomes, peroxisomes, mitochondria, the endoplasmic reticulum and the Golgi apparatus. Vesicles are small membrane enclosed spheres that mediate the traffic of membrane lipids and of water-soluble contents between cellular compartments. The main feature of lipid membranes is their impermeability to water and water-soluble metabolites. Only through proteins, such as transporters or channels can ions and water-soluble metabolites cross the membrane. Biological membranes are comprised of a bilayer of lipids. This is a consequence of their physical properties. Similar to olive oil forming a droplet in water, membrane lipids form droplets in water due to their water repelling nature (also called hydrophobicity). It is energetically favourable for lipids to stick to each other and for water molecules to stick to each other (hydrophobic effect). In this arrangement water molecules can form hydrogen bonds with each other, and lipids can form weak bonds through van der Waals forces. In contrast to olive oil, membrane lipids also have a polar end, which is called the head-group (Fig. 8D: 1). As a result, membrane lipids are amphiphilic (having a polar and a non-polar side) because they interact with water through polar head-groups (see Fig. 8) and aggregate with their hydrophobic fatty acids (Fig. 8D: 2) to form the membrane. Some common lipids are shown in Figure 8 and 9.

The amphiphilic structure of membrane lipids causes them to form bilayers, because it is energetically favourable for the hydrophobic parts to be in contact with each other as shown in the molecular dyanmics simulation video below. The video also demonstrates that a membrane is highly fluidic.

Nervous messages are invariably associated with an electrical change known as the action potential. This potential is generally believed to arise at a membrane which is situated between the axoplasm and the external medium. If this theory is correct, it should be possible to record the action potential between an electrode inside a nerve fibre and the conducting fluid outside it. Most nerve fibres are too small for this to be tested directly, but we have recently succeeded in inserting micro-electrodes into the giant axons of squids (Loligo forbesi).

Sir Alan Hodgkin with Andrew Aelding Huxley, British physiologists recognising the membrane as an electrical insulator in 1939.

Designer

There are many types of lipids in cells. They are most readily grouped by the molecule to which fatty acids and polar groups can be attached (Fig. 9), namely glycerolipids (glycerol backbone), sphingolipids (sphingosine backbone) and cholesterol (sterol backbone).

Glycerolipids can have a variety of head-groups, giving the lipid different biophysical and biochemical properties (Table 3).

Headgroup	Structure	Property	Function
Phosphoryl-ethanolamine	-PO4-CH2-CH2-NH3+	Basic	Membrane structure
Phosphoryl-choline	-PO4-CH2-CH2-N(CH3)3+	Basic	Membrane structure
Phosphoryl-glycerol	-PO4-CH2-CH(OH)-CH2OH	Neutral	Membrane structure
Phosphatidyl-inositol	-PO4-CH6H11O6	Neutral	Signalling
Phosphoryl-serine	H3N+-CH(CH2-PO4)-COO-	Zwitterionic	Signalling

Alternatively lipids are classified by their chemical properties. Glycolipids are a large group of sugar-containing lipids. They often make use of sphingosine as a backbone (Fig. 10) and instead of phosphoryl-choline have complex oligosaccharides attached to the hydroxyl-group of sphingosine. Quantitatively, lipids with a phosphoryl-group linking the backbone to the head-group are dominating. Collectively these are called phospholipids.

Cholesterol (sterol lipid) has some properties that sets it apart from glycerolipids and sphingolipids. The sterol ring system makes cholesterol more rigid compared to the flexibility of fatty acid hydrocarbon chains. As a result it increases the order in lipid membranes and higher levels of cholesterol protect membranes against dispersion by detergents. This is important in the liver where bile acids are produced, which are used to disperse fat, but could also damage membranes. The polar head-group of cholesterol is very small, just -OH, but it ensures its orientation in the membrane. It is however easier for cholesterol to flip from one membrane leaflet to the other, a movement that is near impossible for glycero- and sphingo-lipids. Cholesterol can form esters with fatty acids, rendering the molecule very hydrophobic. In this form it is incorporated into lipoproteins (see chapter 13).

Carbohydrates

Carbohydrates are typically viewed as energy metabolites, but are also important building blocks. The surface of all cells is coated with complex carbohydrate-chains that are attached to proteins (N-glycosylation attached to glutamine and asparagine residues, O-glycosylation attached to serine, threonine and tyrosine residues) and glycolipids (sphingolipids with complex carbohydrate-chains). Carbohydrates are particularly prevalent in proteoglycans, which are extracellular proteins with large and complex carbohydrate-chains attached to them (Fig. 10).

Proteoglycans have modified carbohydrates attached to them, containing amino groups, carboxyl-groups, sulfate groups etc. This allows holding more ions and water molecules within the extracellular space. This provides elasticity for example to our skin. You can squeeze your skin and feel the elasticity of the extracellular matrix. Proteoglycans are found in higher density in bone and cartilage. Another important role of carbohydrates is to provide visibility to the immune system. All extracellular proteins are coated with specific carbohydrates. Moreover, 2-Deoxyribose and Ribose form a building block in DNA and RNA, respectively.

Uric acid (Fig. 11) is often elevated in blood of people with metabolic syndrome (an early stage of type II diabetes). It has low solubility in water.

Answer the following questions:

Q1: Do you think it is a strong acid? Give a reason.

Q2: Compare uric acid to building blocks in the body, from what molecule might it be derived from?

Q3: What problem might the low solubility cause when there is too much uric acid (answer not covered in the text)?

Show answer 1

It is not a strong acid. A strong acid is characterised by complete dissociation. The image shows two different tautomeres in equilibrium.Thus some of the acid is in the ketoform where it cannot dissociate.

Show answer 2

Urica acid looks similar to nucleobases. Our body has no metabolic pathway to completely breakdown nucleobases. Excess nucleobases is released with the urine.

Show answer 3

Low solubility can cause precipitates to form. This can occur in the kidneys generating kidney stones, but more typically in joints causing gout.

Q4. Match the following lipids to their class. Write down and toggle to class. Choices: Sterol lipid, Glycerolipid, Sphingolipid, Polar headgroup, Fatty acid.

Lipid

Class

Lipid

Class

Q5. Which structure of leucine is the most prevalent at pH 7.5?

Show answer

The structure in the center.

Q6. Assign each amino acid to the group with matching biophysical properties. (Choices: anionic, non-polar neutral, polar neutral, cationic, aromatic).

Amino acid

Group