Chargaff's Rules - Research

Research

The second of Chargaff's rules (or "Chargaff's second parity rule") is that the composition of DNA varies from one species to another; in particular in the relative amounts of A, G, T, and C bases. Such evidence of molecular diversity, which had been presumed absent from DNA, made DNA a more credible candidate for the genetic material than protein.

In 2006, it was shown that this rule applies to four of the five types of double stranded genomes; specifically it applies to the eukaryotic chromosomes, the bacterial chromosomes, the double stranded DNA viral genomes, and the archeal chromosomes. It does not apply to organellar genomes (mitochondria and plastids) smaller than ~20-30 kbp, nor does it apply to single stranded DNA (viral) genomes or any type of RNA genome. The basis for this rule is still under investigation, although genome size may play a role.

The rule itself has consequences. In most bacterial genomes (which are generally 80-90% coding) genes are arranged in such a fashion that approximately 50% of the coding sequence lies on either strand. Wacław Szybalski, in the 1960s, showed that in bacteriophage coding sequences purines (A and G) exceed pyrimidines (C and T). This rule has since been confirmed in other organisms and should probably be now termed "Szybalski's rule". While Szybalski's rule generally holds, exceptions are known to exist. The biological basis for Szybalski's rule, like Chargaff's, is not yet known.

The combined effect of Chargaff's second rule and Szybalski's rule can be seen in bacterial genomes where the coding sequences are not equally distributed. The genetic code has 64 codons of which 3 function as termination codons: there are only 20 amino acids normally present in proteins. (There are two uncommon amino acids—selenocysteine and pyrrolysine—found in a limited number of proteins and encoded by the stop codons—TGA and TAG respectively.) The mismatch between the number of codons and amino acids allows several codons to code for a single amino acid - such codons normally differ only at the third codon base position.

Multivariate statistical analysis of codon use within genomes with unequal quantities of coding sequences on the two strands has shown that codon use in the third position depends on the strand on which the gene is located. This seems likely to be the result of Szybalski's and Chargaff's rules. Because of the asymmetry in pyrimidine and purine use in coding sequences, the strand with the greater coding content will tend to have the greater number of purine bases (Szybalski's rule). Because the number of purine bases will, to a very good approximation, equal the number of their complementary pyrimidines within the same strand and, because the coding sequences occupy 80-90% of the strand, there appears to be (1) a selective pressure on the third base to minimize the number of purine bases in the strand with the greater coding content; and (2) that this pressure is proportional to the mismatch in the length of the coding sequences between the two strands.

The origin of the deviation from Chargaff's rule in the organelles has been suggested to be a consequence of the mechanism of replication. During replication the DNA strands separate. In single stranded DNA, cytosine spontaneously slowly deaminates to adenosine (a C to A transversion). The longer the strands are separated the greater the quantity of deamination. For reasons that are not yet clear the strands tend to exist longer in single form in mitochondria than in chromsomal DNA. This process tends to yield one strand that is enriched in guanine (G) and thymine (T) with its complement enriched in cytosine (C) and adenosine (A), and this process may have given rise to the deviations found in the mitochondria.

Chargaff's second rule appears to be the consequence of a more complex parity rule: within a single strand of DNA any oligonucleotide is present in equal numbers to its reverse complementary nucleotide. Because of the computational requirements this has not been verified in all genomes for all oligonucleotides. It has been verified for triplet oligonucleotides for a large data set. Albrecht-Buehler has suggested that this rule is the consequence of genomes evolving by a process of inversion and transposition. This process does not appear to have acted on the mitochondrial genomes. Chargaff's second parity rule appears to be extended from the nucleotide-level to populations of codon triplets, in the case of whole single-stranded Human genome DNA. A kind of "codon-level second Chargaff's parity rule" is proposed as follows:

Codon populations where 1st base position is T are identical to codon populations where 3rd base position is A: « % codons Twx ~ % codons yzA » (where Twx and yzA are mirror codons i.e. TCG and CGA). Codon populations where 1st base position is C are identical to codon populations where 3rd base position is G: « % codons Cwx ~ % codons yzG » (where Cwx and yzG are mirror codons i.e. CTA and TAG). Codon populations where 2nd base position is T are identical to codon populations where 2nd base position is A: « % codons wTx ~ % codons yAz » (where wTx and yAz are mirror codons i.e. CTG and CAG). Codon populations where 2nd base position is C are identical to codon populations where 2nd base position is G: « % codons wCx ~ % codons yGz » (where wCx and yGz are mirror codons i.e. TCT and AGA). Codon populations where 3rd base position is T are identical to codon populations where 1st base position is A: « % codons wxT ~ % codons Ayz » (where wxT and Ayz are mirror codons i.e. CTT and AAG). Codon populations where 3rd base position is C are identical to codon populations where 1st base position is G: « % codons wxC ~ % codons Gyz » (where wxC and Gyz are mirror codons i.e. GGC and GCC). Examples — computing whole human genome using the first codons reading frame provides: 36530115 TTT and 36381293 AAA (ratio % = 1.00409). 2087242 TCG and 2085226 CGA (ratio % = 1.00096), etc...