Friday, September 25, 2009

DNA Replication

Before Watson and Crick suggested model for DNA, it was known that DNA undergoes self duplication or replication but there was no idea how it takes place. One of the major achievement of Watson and Crick model is that it offered basis for DNA replication.

Mechanism of Replication

Watson and Crick proposed a possible mechanism of replication based on their model. Their observation stemmed from the property of complementarity.

According to them:

(a) The hydrogen bonds between the bases of two complementary chains dissolve and the two chains unwind.

(b) Each strand maintains its integrity in the process and does not break down.

(c) Each strand acts as a template (a guide or blue print) or pattern for assembly of another strand, one which is complementary to it.

(d) The complementary strands are constructed from the building material of the nucleotides present in the cell.

(e) As the sugar phosphate back bone is assembled each nitrogen base in the original strand attracts the complementary one. The purine adenine (A) in one chain attracts the Pyrimidine (T).

Similarly T in the other original chain attracts the free units of A present in the cell.

(f) This complementary attraction by all the bases in each of the two original strands along with the assembly of sugar phosphate back bone results in the construction of two new chains, each complementary to the original old ones.

(g) The enzyme DNA polymerase catalyzes the process of replication.

Schemes for DNA replication

There are three schemes.

(1) Semi conservative replication: The above mentioned mechanism for DNA molecule replication is called semi conservative replication because the entire double helix does not remain intact when new DNA is being formed, instead two strands come apart. However each strand is preserved intact. Every daughter DNA molecule has an intact template strand and an intact newly replicated strand.

(2) Conservative Replication: It is alternative method of replication. In this type of replication the original double helix stay together and a new double stranded molecule is built up next to them. The original double helix acts as template for a new one; one daughter molecule would consist of the original parent DNA and the other daughter would be totally new DNA.

(3) Dispersive Replication: The original strands breaks down end entirely new strands are constructed from these and other in the cell. Some parts of original double helix are conserved and some parts are not. Daughter molecules would consist of part template and part newly synthesized DNA.

EXPERIMENTAL EVIDENCE OF REPLICATION

Meselson and Stahl in 1958 provided a strong support for the concept of replication as suggested by Watson and Crick. They designed an experiment to determine the mode of DNA replication. They grew E. coli in medium containing a heavy isotope of nitrogen 15N (normal form of nitrogen is 14N). After growing for several generations on the 15N medium the DNA of E. coli becomes denser (heavy). They employed a technique called density gradient – centrifugation to determine the density of DNA strand. When extracted from cells this heavy DNA settled out in the density of DNA strands. When extracted from cells this heavy DNA settled out in the density gradient at characteristic position.

Replication is bidirectional: Carins established that replication initiates at a single site and it is bidirectional i.e. two replicating sites move in opposite directions around the circular DNA. He also determined the time of movement of this work around the circular DNA.

Replication in Eukaryotes: In Eukaryotes DNA molecules (chromosomes) are larger than in Prokaryotes and are not circular.

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