Part XX: DNA or Protein; The Great Debate

The story of the true carrier of genetic information from generation to generation is an interesting tale which followed a winding path from the late 1800’s up to 1953 and even that historic discovery, while one of the most sought after and important discoveries of human kind, was really just a new chapter in the book of life as we know it that continues today and undoubtable will as long as human civilization exists.

The story really begins in 1869 when a German physician Fredrich Miescher discovered a white, sugary, slightly acidic substance that contained phosphorus.  He named it “nuclein” based on its presence only in the nucleus of cells.  Fast forward to 1914the year that Prince Ferdinand of Austria was assassinated thus beginning “The Great War” (World War I) Robert Fuelgen, another German, found that DNA was strongly attracted to fuchsin, a red dye.  Just a small step but helpful one.  Later it was discovered that DNA was present in all cells and was associated with the chromosomes which had only recently been discovered.  Let’s take a small “aside” now and delve a little further into the world of the chromosome.  Oscar Hertwig, yet another German (an embryologist) observed sea urchin fertilization and realized that only one sperm cell was necessary to fertilize an egg and further noted that when the sperm cell penetrated egg the nuclei from the sperm and egg fuse.  This established that the nucleus, or something in it, is the carrier of genetic information from generation to generation, another small step.  At about the same time, Walther Flemming observed the “dance of the chromosomes” during cell reproduction, “steps” of mitosis (cell division), and the events of each step. Of course, in reality, the process is continuous and people devised the “steps” to understand the process better.  To learn more including the phases such as prophase, metaphase, anaphase, telophase, etc., simple Google “mitosis” or “cell division” or consult any high school biology textbook.    In one experiment Flemming removed the nucleus of an ameba (amoeba for our British friends) and watched the cell die. 

P.A. Levene, a biochemist proved in the 1920’s that DNA was composed of a 5- carbon sugar (a pentose); a phosphate group, and four nitrogenous bases, adenine and guanine (purines) and thymine and cytosine (pyrimidines).  He concluded that each nitrogenous base is connected to a sugar molecule which is attached to a phosphate group which makes up a nucleotide.  (See essay IX Majors)

In 1928, Frederich Griffith, an English bacteriologist public health official was trying to develop a vaccine against Streptococcus pneumonia which causes a form of pneumonia.  This bacterium comes in two forms, one a virulent (disease causing) form with a polysaccharide (simple sugar capsule and a nonvirulent (harmless) none capsulated form.  Griffith wanted to know if injections of heat killed virulent pneumoiae could be used to immunize against pneumonia.  At one point he injected mice simultaneously with heat kicked virulent bacteria and living non-virulent bacteria expecting the ice to live but all the mice died—a complete surprise.  Autopsies on the mice revealed that their bodies were filled with living encapsulated virulent bacteria.  Years later it was shown that extracts from heat killed virulent bacteria when added to harmless bacteria could transform them into harmful bacteria complete with protective capsules.  Thus an extremely important phenomenon called transformation was discovered and the yet unknown substance responsible for this transformation was called a transforming factor. (“Factor” is still used to refer to an unknown substance).  Later this transforming factor was identified as—you guessed it—DNA.  When I presented this in class I usually describe the heat killed bacteria as never-say-die KILLER BACTERIA for dramatic effect.  It was an American scientist, O. T. Avery that identified DNA as the transforming factor.

At this point it must seem like          DNA was the overwhelming candidate for carrying the genetic information from generation to generation—but, hold on, don’t count the other contender out yet.  Max Delbruck and Salvador Luria two scientists who emigrated from Europe during the intellectual mass exodus of the 1930’s along with Albert Einstein,       

                          And other mathematicians and physicists prior to the Third Reich takeover of 1939performed some Nobel Prize experiments in 1940 with a special group of viruses called bacteriophages or simply phages because they infect bacteria.  Yes, even bacteria have enemies.  The particular viruses of interest in their research ae called coliphage and they attach Escherichia coli.  They were numbered T1 through t7 (“T” means “type”). In a personal note here I very recently inoculated E. coli bacteria with T4 coliphage in my home lab attempting to do a T4 assay, See the photo below.

Skipping the details of the experiments, chemical analysis of fragments of phages after infection revealed what we all know now – that viruses are composed of just two organic compounds, a DNA core and  a protein coat. Later RNA was also discovered in the core but if so DNA is absent (see essay IX).  So now the great debate ha had been brewing for years suddenly intensified.  What carried the all-important genetic code, DNA or protein?  Scientists were divided into two camps.  The protein backers had a very simple philosophy:  DNA is composed of just 4 nucleotides but proteins are composed of 20 amino acids (see essay IX).  They reasoned that like an alphabet composed of 20 letters could spell more words than one with 4 letters, so proteins could account for greater genetic variability (diversity).

The stage was now set for the most compelling evidence yet.  In 1952 two researchers Alfred Hershey and Martha Chase performed a brilliant set of experiments based on two simple differences in DNA and proteins.  But first I always made a point of emphasizing that like Rosiland Franklin, Martha Chase was a pioneer in that here was a woman that rose to prominence in a field dominated by men and they could do the same much to their delight.  OK, another personal aside here.  

A few years ago (2013), I ran into a former student in a local Kohl’s Department Store.  She also became a lab assistant and a very good one who earned her degree at the University of Iowa and now was doing scientific research at Marquette University. She told me that she got her start after being my student and lab assistant.  Talk about delight!  I was on cloud 9 at a time I was going through a long, tough, tough time medically.  Then she topped it off by saying that I looked the same as I did when I was her teacher in the mid ‘80’s.  This was at a time when I looked terrible and felt much worse.

Back to Hershey and Chase:  DNA as we know, contain phosphorus but proteins don’t.  Proteins contain sulfur but DNA doesn’t.  They prepared two groups of viruses, one which was labeled with radioactive phosphorus (³²P)and one labeled with radioactive sulfur (³⁵S) and inoculated then into an E. coli host with the appropriate radioactive isotope.  Again skipping some of the details one culture of bacteria was infected with ³²P and another infected with ³⁵Sphage and later were tested for radioactivity.  Their brilliantly conceived experiments revealed that the ³⁵S phages had remained outside the bacterial cells but the ³²P (DNA) had entered the cells, infected the bacteria and produced new viruses.  The great debate was essentially over; DNA was proclaimed the winner.

Conclusion

Aftermath

 Erwin Chargaff of Columbia University analyzed the purine and pyrimidines content from many different species of organisms.  Here is a sample of his results showing percentages of the four bases.

                                           Purines                                            Pyrimidines
source                   adenine                 guanine                 cytosine                thymine

Human                  30.4%                   19.6%                   19.9%                   30.1%
Ox                        29.0                      21.2                      21.1                      28.7

Wheat germ          28.1                      21.8                      22.7                      27.4

after examining the data what can you conclude?  Answer as usual at the end of the essay.

It is almost anticlimactic to now talk about James Watson and Francis Crick’s famous discovery in 1953.  Watson, a former Whiz Kid from Chicago was going to become an ornithologist but thankfully changed careers.  Francis Crick, a trained physicist were an unlikely fil for one of the most important discoveries in the history of science.  Rather than swell on their work which would extend this very long essay much longer I would recommend reading a form of Watson’s famous book “The Double Helix” and again read the last p [art of essay IX (majors).  I have read his book at least three times and learned more each time.  It’s written so that most ordinary people can understand it with just a little science background.  If you can/t or don’t want to read the book (a short one in terms of book length, I would suggest Googling “The Double Helix”.

I briefly described protein synthasis in essay IX but now direct you to a “code of life
chart”.  One of the main mysteries to be delved was “breaking the code”, that is, to learn how

DNA directs:

1. its own replication
2.  protein synthesis

Now that we know DNA directs protein synthesis let’s look in more detail at the overall process.  According to “central dogma”, a term that Crick himself coined, during protein synthesis the double stranded DNA molecule splits down the middle (hydrogen bonds break releasing emery) and each side serves as a template for a strand of messenger RNA (mRNA with the cytocine of DNA coding for a guanine of mRNA and adenine of DNA coding for uracil of mRNA.     Remember that thymine in DNA is replaced by uracil in RNA. However, thymine of DNA still codes for adenine of RNA.  This whole process is called transcription, a process that occurs in the nucleus.  The single stranded mRNA leaves the nucleus and pairs with a transfer RNA (tRNA) which contains an amino acid (the building block of proteins) on one end and an attachment on the other end to enter a ribosome (ribosomal RNA or rRNA).  As stated in an earlier essay, it’s as if the mRNA molecule says to the tRNA, “let’ meet at the ribosome (more correctly let’s go to the ribosome) and make a protein”, a process called translation. The long strand of mRNA can be thought of as individual units of codons (three nitrogenous bases per codon) that like transcription from DNA, pairs with the corresponding anticodon on the tRNA molecule. That is, a u from mRNA pairs with an a of tRNA, c with g,  g with c, etc.  Remember though an a pairs with a u of tRNA. Why? The individual tRNA subunits join together inside the ribosome (rRNA) and exit out in long polypeptide  chains which when long enough and exhibit secondary structures and perhaps tertiary and quaternary structures are called proteins.  But remember, and this is huge, it all began with DNA.

Let’s pause for a moment and reflect on some important principles.

1. DNA is composed of 4 nucleotides, a four letter alphabet (the four nitrogenous bases)
2. There are 20 amino acids commonly found in living organisms
3. Codons (and, therefore, anticodons) occur in triplets

Now the question arises of why groups of 3 bases in a codon?  Why not 1 or 2?

Answer:

• Individual codons could code for only 4 amino acids. (4¹) 4 bases raised to fist power if just one individual base
• Pairs of codons could produce only 16 amino acids (4²)
• Triplets could produce 64 amino acids (4³) 4x4x4= 64 which is more than enough for the 20 amino acids  In fact, this means that there must be more than one codon that can code for each amino acid, right? Right.

Now we are ready for the ultimate task and the climax of this entire essay and perhaps all of my essays except for perhaps those on climate change.  Identifying the correct structure of the DNA molecule is one thing but to apply it to genetics and understand how I works in specifying the production of the thousands of chemical compounds and chemical reaction each with its own enzyme in our bodies is a totally different set of circumstances.  But let’ proceed.  Here is the “code of life” chart that summarizes (I didn’t say explains) all of the previous sentences.

The letters along the left side, top, and right side represent the 4 bases. You know their names by now.  The letters (in triplet) represent the codons and the abbreviation of individual amino acids. To select amino acids:

1. choose a letter on the left side
2. choose a letter on top
3. choose a letter on the right side
4. write them down and identify the amino acid using the listing below using the abbreviations.

Credit to: RearchGate

I’ll do two examples for you.

Q.  Find the code(s) for cysteine (cys)

A.  UGU, UGC

Q.  Find codes for serine (ser)

A.  AGU, AGC

Let’s reverse the process. 

Q.  What does the codon CCU code for?

A.     Arginine

Now for a real problem.

Find the code for

          Alanine(ala)                   plus glycine(gly)            plus valine (val)

Feeling pretty good? Try another code for them.