Chapter 3

I am filled with wonder that we live in an era when we can consider the world in which we live and the physical nature of our solar system with the added ability to verify what we see.

The territories of human learning are named and categorized.

Astronomy is now a major human science. (Astron=star and nomos = law)

We know that the earth has a diameter of 12,756 kilometres, and that it is rotating around the sun, which is about 150 million kilometers away from us.

Since the earth's orbit is elliptical, this distance must vary.

The sun itself has a diameter about 1,392,000 kilometres.

But we also know that the whole solar system is about 30,000 light years from the centre of a spiral galaxy which we call the Milky Way.

(Light travels at about 300,000 km per second in a vacuum, which comes to about 9.5 trillion km in one year)

This galaxy contains about 300 billion stars, but is only one of billions of galaxies in the Universe.

In the Monty Python comedy film "The Meaning of Life"the song about the universe tells us that we go around this centre , taking 250 million years to do so!

In our small part of this universe life has appeared, and advanced into complexity.

All living things are composed of elements and those that connect together in living things make up the many building blocks of carbohydrates, fats (lipids) and proteins, together with essential compounds called vitamins.

Scientists have found it convenient to have specific names for the molecules of life.

I want to introduce some of these concepts to the non-biologist, particularly because science is increasingly able to explain more and more about the inherent properties of molecules, which make the march of evolution into more complex life possible.

As I write I am conscious that what I am doing is one of many ways of representing our structures and functions to our selves and to others.

Hundreds of volumes of writing are required to even describe what is known so far.

It is usual rather than unusual for specific biological scientists to know their own areas but have major gaps in their knowledge of other molecular biological territories.

A return to basics is a required step when we are to integrate what is being uncovered with the texts of the recent past.

Those of you who have studied chemistry will have some exposure to these concepts or representations.

In some rather mysterious way, when we look again and again at certain forms or representations, they become familiar to us.

Someone has said, "Whatever we do a lot of, we become good at"

How important can it be to recognize our own patterns?

We now address both parts and wholes, and the patterns that enable them to connect.

This is the mystery of life.

Everything that has unfolded in the history of the Kosmos is grounded or founded upon the capacity of atoms and molecules to be arranged with emergent properties.

It is a dynamic happening, which is difficult to capture in prose.

Hydrogen

This simplest of all elements is the most plentiful atom in the Kosmos.

It consists of one proton as the nucleus and one electron in orbit around the proton.

 

 

Water

Water itself is an amazing substance, with electrons changing more that 1 billion times per second in the bonds between the 2 hydrogen atoms and the oxygen atom. (H20)

It is described as a very polar molecule. The oxygen has a strong negative charge, and the hydrogens have positive charges

Molecules that dissolve in water are able to do so by being able to dissociate into positive and negative ions (e.g. Salt is sodium chloride'Na+ and Cl-)

Hydroxyl (OH) groups on glucose are polar, and make glucose almost infinitely soluble in water.

Science maps the possible and can confirm whether molecules can interact and if so, in which ways.

There are also ways to number the atoms and designate where a group is located.

Carbohydrates

These have the formula Cx (H20) y, sometimes with attached other pieces.

The smallest carbohydrate units are called monosaccharides (sugars) with longer groupings (two= disaccharides, 3-11= oligosaccharides and longer chains being called polysaccharides)

The general terms are 1= monomer, 2= dimer, 3= trimer, increasing to oligomers and polymers)

Plants use the process we call photosynthesis to combine carbon dioxide and water into carbohydrates and giving off oxygen.

Sugars may contain amino groups (e.g. glucosamine and galactosamine)
The amino groups are often acetylated.

Inside cells monosaccharides usually contains phosphate groups. This results in the phosphorylated sugar being unable to cross membranes.

Phosphate groups may join sugars to nucleosides.

Only after a close look at the process of phosphorylation,does one realize the amazing number of biological reactions which involve phosphorylations.

Some of these will be described as we explain mechanisms in chapters to come.

Sulphated sugars are found in connective tissue.

Glycosides are formed when the hydroxyl group on the anomeric carbon of a monosaccharide react with the -OH or -NH of another compound.

These bonds are called glycosidic or glycosyl groups.
N-glycosidic groups are found in nucleotides. (e.g. In adenosine triphosphate (ATP), the base adenine is linked to the sugar ribose via an N-glycosidic bond.)

The reader can note the forms of substances that are vital to certain life happenings.
In this example, ATP is storage energy for cell activities

Fats

Fats (oils and lipids) are made up of fatty acids, esterified fatty acids (e.g. glycerol), and interesting acyl glycerols (here fatty acids react with alcohol (hydroxyl groups). Triacylglycerols are called triglycerides.

They are called hydrophobic in that they are not very soluble in water.

The fatty acids are chains of carbon atoms with a carboxyl (COOH) group at one end and a methyl group (CH3) at the other end. (This latter is called the N or omega ( ) end)

Short fatty acid have chain lengths of 1-4 carbon atoms, medium chains are of 4-12 carbon lengths and long chain 14-20. (There are longer chain fatty acids sometimes called very long chain fatty acids>20 carbons.

If there are no double bonds in the molecules they are called saturated fatty acids.

If there is a single double bond(C=C) in the molecule the fatty acid is called a monounsaturated fatty acid, and if more than one(C=C-C=C), a polyunsaturated fatty acid.

Most fatty acids in human beings are even numbered and are usually16-20 carbon lengths.

It is now known that in all biological membranes, the basic permeability barrier is a lipid bilayer (double layer)

It was discovered that the lipid molecules and the protein components of these membranes are free to exhibit a variety of motional modes, such as translation, vibration and rotation which endow these membranes with dynamics such as are needed for the crossing of the membranes by crucial molecules of life.

In biological membranes there are three classes of lipids, namely

Glycero-phospholipids, sphingolipids, and sterols.

The arrangement in certain cell locations exhibits considerable diversity.

Transport of fatty acids into cell membranes can be important, as for example long chain fatty acids(C 20 and higher) need acetyl carnitine to transport them into mitochondrial membranes.

With the existence of this lipid layer membrane, there is a barrier that nature has used to evolve localized structures to enable cells to sense and respond to changes outside themselves.

Cells may respond to an amazing array of stimuli, including amino acids and peptide structures, products of cell metabolism, ions, and even photons.

In the writing ahead I will refer to these recognition units or receptors, along with coupling structures and gene responses to these signals.

Gene products are made and chemically changed by other gene products in order to carry out crucial functions.

These responses are often specific, but result in other intracellular events.

It would be a scintillating sight if we could see what is happening in cells in every second of their lives!

For readers who want to study the basic structures and functions of lipids, I direct them to any recent biochemistry textbook, and I will mention the optimal dietary requirements in the section on therapy.

Medical biochemists may be tempted to study only human biochemistry, but in truth we need to grasp the chemistry of all living things and the domains of their existence.

Proteins.

Aminoacids are the monomeric units from which proteins are assembled.

Two amino acids combined= dipeptide, 3-5= oligopeptide, and longer chains are called polypeptides.

Depending on the chemical groups on the amino acids (great diversity here), the amino acid may be nonpolar, polar uncharged, or polar charged.

The side chains are crucial to specific amino acid functions.

Amino acids that our bodies cannot make are called "essential amino acids".

Fascinating new molecules with diverse functions emerge when carbohydrates are combined with proteins (glyco-proteins), as well as when lipids are combined with proteins (lipo-proteins)

Enzymes are proteins which increase the rate of chemical reactions.

We call the chemicals acted upon by enzymes "substrates", and the resulting compounds from these actions are "products".

The balance of these reactions in living systems can be measured.

Isoenzymes are variants of an enzyme with different aminoacid sequences and different properties.

Genes.

Genes are of course the units of heredity.

Germ cells (ova and spermatozoa) carry mostly the genes of ancestral lines, but as well mutations, some of which are virally and environmentally induced

Genes are constituted by sequences of bases called adenine (A) and guanine (G) (purines) and cytosine(C) and thymine (T)(pyrimidines), which are linked to sugars and phosphates as nucleotides.

A nucleoside consists only of the base plus the sugar, and when the phosphate is added it is called a nucleotide.

The genetic information is coded in DNA (desoxyribonucleic acid).

The only way the bases can pair is T-A and C-G (Watson and Crick) and the sequences form the famous double helix, which these researchers described.

Most of the DNA in our cells (eukaryotic cells) is in the nuclei, but some viruses have DNA, which they are unable to use unless they are inside cells.

Viruses may have between 20 and 200 genes, while prokaryotes (bacteria) have1.000-3, 000 genes.

The nematode worm, Caenorhabditis elegans has about 19,000 protein coding genes, while human beings appear to have about 25,000 genes.

In the following discussion about non-protein coding genetic material, I will attempt to give insights into recent discoveries about "non-coding" DNA and surprising new information about RNAs.

It is difficult to explain viruses without seeing them as products of prokaryotic (and perhaps later eukaryotic) cells.

The DNA carries the master codes from which the body can synthesize proteins.

RNA (ribonucleic acid) has the base uracil connected to the adenine rather than thymine, U-A) and its sugar is ribose.

In eukaryotic cells (cells with nuclei), messenger RNA (mRNA) is transcribed from DNA in the nucleus, and travels through pores in the nuclear membrane to the cytoplasm.

This transcription is a copy of the totality of the gene as an RNA transcript.

Then follows a process called "splicing", where intronic RNAs are removed leaving the coding sequence we call messenger RNA.

Transfer RNA (tRNA) molecules carry amino acids to ribosomes, which are the structures in which proteins are assembled through the messenger RNA.

Ribosomes can also migrate from nucleus to cytoplasm through the pores.

Ribosomal RNA (rRNA) can vary in type from prokaryote (cells without nuclei such as bacteria) to eukaryote and combine with proteins to form ribosomal subunits.

Prokaryotic ribosomal subunits are called 70S and have subdivisions 50S (about 34 proteins) and 30S (about 21 proteins). These will be mentioned when I describe certain antibiotic actions.

Eukaryotic (cells with nuclei) ribosomal subunits are called 60S (about 50 proteins) and 40S (about33 proteins)

In essence these names or codings are the invention of scientists who discover ribosomal structures and their functions.

Other RNAs are involved in primers for DNA replication and other splicing and modification reactions in RNA precursor assembly.

RNA also serves as the genome for certain viruses (eg HIV)

In general the sequence of information is from DNA to RNA, but in retroviruses like HIV, an enzyme called reverse transcriptase uses the RNA genome to produce a DNA copy.

The human genome project, together with intricate details of the functional molecular consequences, is truly revolutionizing biological knowledge.

We have paid much attention to genes as carriers of codes for protein sequences, but so-called coding DNA accounts for less than 1% of nuclear DNA in human beings.

I would like the reader to know more about this vast amount of human DNa which codes for RNA, but not for protein synthesis.

Non-coding DNA

DNA sometimes codes for RNA sequences that are not translated.

Names are given to specific regions along chromosomes.

Exons are portions of genes (DNA sequences) that are eventually spliced together to allow formation of messenger RNA, and code for protein assembly.

Introns are segments spliced from precursor RNAs during RNA processing located in spacing between exons. They do not code for proteins.

Research has revealed regulators of genes in at least three locations.
(1) Promoters, which are upstream of transcription start sites,
(2) Inside introns
(3) Downstream, in a polyadenylation tail region)

Most genes have at least 15-20 discrete regulatory elements within 300 base pairs of transcription start points.

Perhaps as many as35-50, 000 RNA only genes are located within non-coding DNA regions.

Some1,194 sequences are highly conserved, showing very little variation between rats, dogs, cows and human beings. Of these about two- thirds were in introns, 244 in coding DNA and the rest in noncoding DNA.

The name "pseudogenes" refers to non-protein making genes, which are complementary DNA sequences on the other side of the DNA ladder.

In some cases the alter-ego churns out antisense DNA.

If sense and antisense RNAs meet, the gene cannot express the protein.

The example is Makorin1 p1, which is a pseudogene copy of makorin1, and cannot make the protein, but when it is knocked out, makorin 1 shuts down.

It may well emerge that as many plants and bacteria as well as >1,600 human genes can express anti sense RNA, there is a big potential for the interplays to bring about both defensive functions, but also pathogenic consequences.

By 2003, some 150 micro RNAs have been found in human beings. Many more will be found.

Nelson Lau and David Bartel write of the discovery of RNA interference (RNA-i), which can silence expression of threatening genes, by interrupting only the offender's messenger RNA without disturbing the messages of other genes.

Viruses on entering cells may activate a blocking effect through RNA-i

Andrew Fire and Craig Mello observed potent silencing effects on the unc-22 gene of C elegans when the worms were inoculated with corresponding unc-22 double stranded RNA, when neither the corresponding single-stranded RNAs, whether sense or antisense, had any effect.

There are genes to convert single-stranded RNA into double stranded RNA.

This was illustrated by Douglas at Oregon State University, in the tobacco etch virus and genetically engineered tobacco plants that contained copies of the coat protein gene of the virus, Some plants proved to be resistant to this virus.

This led to the exploration of the nature and function of silencers.

Enzymes such as PKR can block translations of all messenger RNAs (both normal and viral) and RNAse-L indiscriminately destroys messenger RNAs.

How does RNAi work?

Inside the cell, the double stranded RNA encounters an enzyme dubbed Dicer, which cleaves the long RNA into pieces.

These pieces are 22 nucleotides long and are known as short interfering RNAs (siRNAs)

Dicer cuts through both strands of the long double-stranded RNA at slightly staggered positions, so that each resulting siRNA has two overhanging nucleotides on one strand at either end. The siRNA duplex is unwound and one strand is loaded into an assembly of proteins to form the RNA-induced silencing complex (RISC)

Positioned so that messenger RNAs can contact it, the siRNA of RISC will adhere only to a messenger RNA that closely complements its own nucleotide sequence.

Unlike the interferon response, the silencing complex is highly selective in choosing its target messenger RNAs.

When a matched messenger RNA docks onto siRNA, an enzyme known as Slicer cuts the captured RNA strand into two.

The RISC then releases the 2 mRNA pieces (which are now incapable of directing protein synthesis) and moves on, staying intact and free to find and find and cleave another mRNA.

Tuschl and colleagues put synthetic siRNAs into cultured mammalian cells and demonstrated silencing of target genes with no interferon response.

This knowledge has potential applications in cancer and viral infections.

In normal development genes may be required to be active in embryonic cells and yet be readily turned off later. Yet others need to be turned on later, so RNAi and siRNAs are some among the microRNAs) of different origins) with specific functions as described above.

With hindsight, it seems obvious that cell mechanisms would run into chaos without systems like this.

In summary, RNAs have major roles in catalysing, signalling and switching in genetics.

Mitochondria in cell cytoplasm also contains some DNA (see later)

This is entirely from the maternal line (ova) as sperm mitochondrial DNA is lost in fertilisation.

The science of genetics has advanced beyond our initial understandings.

Professor John Mattick, writing in Scientific American in October 2004, writes that the credo"one gene, one protein", may be largely true for prokaryotes (one celled organisms which lack a nucleus) but must be expanded in eukaryotic organisms which possess nuclei.

He writes, "Proteins do play a role in the regulation of eukaryotic gene expression, but a parallel regulatory system consisting of RNA acting on DNA, RNAs and proteins is also at work."

This signalling network is likely to be crucial to achieve the complexity seen in higher organisms.

It seems likely that the introns are a later occurrence in the evolution of DNA sequences.

Self-splicing mobile genetic pieces are termed group II introns, with properties that allow insertion into genes and abilities to splice themselves out when expressed as RNA.

Group II introns are seldom found in bacteria, but with eukaryotic cells, transcription occurs in the nucleus and translation in the cytoplasm. allowing time for intron RNA to excise itself.

A further development appears to be the emergence of "splice-somes" which are complexes of small catalytic RNAs and many proteins, possessing a capacity to snip intron RNA out of messenger RNA precursors.

In effect this allows intronic RNAs to be involved in their own evolution.

This would explain parallel regulatory systems which need coo-ordination.

RNAs can encode short sequence specific signals doing such tasks as directing RNA molecules to receptive targets in other RNAs and DNA.

The RNA-RNA and RNA-DNA interactions could create structures that recruit proteins to convert signals into actions.

Analogies arise with the handling of bit information in computers.

Feedbacks and feed forward mechanisms are the essence of cybernetics.

To follow development of the fertilized egg to the 700 trillion cells of the adult human being requires such an elegant cybernetic system. involving astonishing co-ordinations of co-ordinations.

Two steps have been studied.

(1) Modification of chromatin

Recent studies suggest that RNA signalling directs the tagging of chromatin determining whether the genes in stretches of DNA will be accessible for translation or stay dormant.

(2) Alternative splicing.

This process generates divergent repertoires of RNAs and proteins in the cells of different tissues.
This is almost certainly RNA controlled, partly by the tagging or grabbing of sequences and partly by directing spliceasome function

The point to note is the capacity for human genes to code for more than one protein, and the number may turn out to be quite large!.

Crucial elements in higher living systems are highly conserved between species.

Something is needed to guide these systems to complexity with minimum errors.

The word "cybernetic" refers to "controlling systems".

The more complex the system, the more sophisticated is the control system.

Mattick points out that if life began on Earth about 3.8 billion years ago, a leap in complexity must have occurred about 1 billion years ago and more so in the Cambrian "explosion" of invertebrates about 525 million years ago.

The genes, therefore, do not sit as isolated pockets in a sea of junk DNA, but rather they live in a regulatory environment, which is utterly crucial for our complexity.

Non-coding RNAs have already been linked to B cell lymphomas, lung cancer, prostate cancer, autism and schizophrenia.

In essence we now understand that our genes, through gene products, and now non protein coding genetic materials, shape the possibilities of our fundamental functions, but as well shape a changing response to environmental influences over varying time scales.

But there is more to the complexity of life than this!

Recent research is revealing amazing changes in gene expression and activity related to environmental happenings, and this will form a basis for particular protections for many diseases.

As mentioned earlier, in specific circumstances genes are activated and cell chemistry changes.

Anyone who studies genetic engineering knows that some viruses can be used to splice genes from one species into another.

Viruses are involved in extensive transfer of DNA and RNA!

It is self evident that scientists are exploiting a natural capability of some viruses.

It should be noted that there are nonviral means for DNA to be transferred from cell to cell.

Intra cellular organelles called plasmids can be transferred between prokaryotes and also be carried into eukaryotic cells by organisms which enter cells.

As long as life exists, there is no possibility of there being no evolution.
Evolution involves such gene insertions, and mutations as well as the direct inheritance of parental or ancestral genes.

Now we can add other mechanisms for inserting functional elements. (RNA editing) and transposons (previously thought to be junk!)

Mattick mentions the discovery of adenosine to inosine (A to I) editing discovered by Erev Levanon and colleagues and announced in July 2004.

Again they arise in non-coding RNA sequences called Alu elements.

A to I editing sequences are very active in the brain and aberrant editing has been associated with epilepsy and depression.

It is not hard to suppose that the elegance of neural function demands huge programming flexibility.

We do well to honour the discovery of genes and we will do better as more details of patterns of gene activation and transcription of codes reveal participation in the web of life!

For the moment, the message is that as long as we are biological beings, each of us can decide to honour our own life, the life of others and the biological economy, which is underpinning our present and future.

Does this make you rethink what short term claims about drugs really mean in our whole living patterns?

The concentrations by business organizations and politicians on monetary policies, and the so -called economic rationalisms will bring great disasters unless we recognize, ,support and nurture the biological economy which underpins all life.

Are the "John Matticks' of this world too rare in a climate of drug company directed research?

We can be encouraged that wonderful discoveries are being made in private genetic companies, yet remain puzzled about an overall balance as to how to justly share information.

Let us love biology.

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john.graham@flinders.edu.au

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