Foundations of a replicative organism

DNA Discovery by Friedrich Miescher (Swiss, 1844-1895)

He discovered a substance containing both phosphorus and nitrogen, made up of molecules that were apparently very large, in the nuclei of white blood cells

Named the substance nuclein because it seemed to come from cell nuclei. In 1874 when Miescher separated it into a protein and an acid molecule. It is now known as deoxyribonucleic acid (DNA)

He worked with Albrecht Kossel and Emil Fischer, the nucleic acid and protein experts at the turn of the 20th. century

Phoebus Levene

He conducted experiments that in 1931 suggested that the four components of DNA

occur in approximately equal ratios

He suggested the possibly that DNA was made of a repeating tetramer

If so, the implication was that the structure of DNA was too simple and too regular to contribute to genetic variation: attention

thereafter focused on protein as the probable hereditary substance

Not only did Levene identify the components of DNA, he also showed that the components were linked together in the order phosphate-sugar-base to form units. He called each of these units a nucleotide, and stated that the DNA molecule consisted of a string of nucleotide units linked together through the phosphate groups, which are the 'backbone' of the molecule.

Scientist thought that Proteins (made from 20 aa) wereNeeded to encode life, not the 4(?) forms of Nucleotides

The Tetranucleotide Hypothesis

Erwin Schrödinger published in 1945 a book titled What is Life? that planted the idea for searching “the secret of life”

Chargaff noted the publication of Avery, MacCleod and McCarty paper and realized that DNA was the key to life and set out to prove Levene wrong

In 1944 Consden et al. showed that it was possible to separate individual amino acids and to determine the amino acid composition of protein hydrolysates by partition chromatography on paper strips. The method was, in principle, readily adapted for the separation and identification of a large number of other substances, includingthe purines and pyrimidines of the nucleic acids(Figure 1), a task carried out in Chargaff’s laboratory by the Swiss post-doctoral fellow Ernst Vischer [12], andindependently at the Rockefeller Institute by Rollin Hotchkiss

Note C not always equal to G

Apart from not demonstrating equal amounts of the fourbases and thus casting doubt on the validity of the tetranucleotidehypothesis, certain other unexpected patternsalso emerged: the amounts of purines seemed always toequal those of pyrimidines (that is, A + G = C + T, or(A + G)/(C + T) = 1). This had been found by Alfred Mirskyin 1943, but seems to have been overlooked by theChargaff laboratory. More curiously, the ratios of A:G andT:C were always similar to each other whether they weregreater or less than 1

The significance of these relationships was puzzling anda constant source of comment. At the end of 1949 Chargaffnoted that ‘‘A comparison of the molar proportions [of thebases] reveals certain striking, but perhaps meaningless,regularities’’. Early in 1950, he wrote ‘‘It is noteworthy,although possibly no more than accidental, that in alldesoxypentose nucleic acids examined thus far the molarratios of total purines to total pyrimidines were not farfrom 1. More should not be read into these figures.’’Later in 1950, apparently as a last-minute insertion in thepaper, Chargaff wrote ‘‘It is noteworthy – whether this ismore than accidental, cannot yet be said – that in alldesoxypentose nucleic acids examined thus far the molarratios of total purines and total pyrimidines, and also ofadenine to thymine and of guanine to cytosine [ratioscuriously not actually presented], were not far from 1’’[2]. The following year, he wrote ‘‘As the number ofexamples of such regularity increases, the question willbecome pertinent whether it is merely accidental orwhether it is an expression of certain structural principlesthat are shared by many desoxypentose nucleic acids,despite far-reaching differences in their individual compositionand the absence of a recognizable periodicity in theirnucleotide sequence’’. He then added ‘‘It is believedthat the time has not yet come to attempt an answer’’,although clearly the subject was very much on his mind.

Why didn’t Chargaff predictThe Double Helix?

-He didn’t want to be wrong like Levene!

Two reasons why he would have been wrong

1)

2) What if it was some other reason thatA/T G/C ratio was the same????

3) H and X nucleotides?

Accounting in Saccharomyces cerevisiae

One of Chargaff’s Rules