The Cell - Challenges in Delivery of Therapeutic Genomics and Proteomics - page 10

developed method that involves visualizing the chromosome in bright colors is called
chromosome painting, wherein a fluorescent probe specific for sites on the chromo-
somes is used [1-2] .
1.2.2 DNA Replication, Repair, and Recombination
It has been seen that the two strands of DNA have complementary base pairing, thus
enabling each to serve as a template for the synthesis of a new complementary strand.
This is the basis of DNA replication and recombination. In eukaryotes, the replication
machinery, a group of proteins, ensures correct and efficient replication of the DNA
strand. The DNA replication is semiconservative in nature, with each double helix
generated by replication containing one parent strand. DNA replication begins with
separation of the two strands at the specific replication origins—regions rich in AT
base pairs—brought about by origin recognition complex and some other proteins like
CDC6, CDT1, and MCMs. The DNA double helix is unwound near the replication ori-
gin, and in this place a Y-shaped structure called a replication fork is generated by new
strand synthesis along the parent strand. A protein called single-strand binding protein
or replication protein A attaches to the unwound DNA strand and prevents reforma-
tion of the base pairs by hydrogen bonding. The enzyme topoisomerase relieves the
stress created from the unwinding of DNA strands. DNA polymerase, the most impor-
tant enzyme of the replication machine, is involved in the synthesis of new strands by
catalyzing the addition of nucleotides at the  end by esterification. However, the poly-
merase requires a primer to which it can add nucleotides. An oligonucleotide RNA/
DNA usually serves as a primer that is synthesized by enzyme DNA polymerase .
Subsequently, enzyme DNA polymerase  moves along the DNA strand as it attaches
more and more nucleotides. The direction of elongation is always in 5→ direction.
Because the two strands are antiparallel, the nucleotide addition cannot proceed con-
tinuously in the same direction on both strands.
The continuous strand is called the leading strand. On the other strand, DNA is
synthesized in small fragments (Okazaki fragments) in 5→ direction discontinu-
ously and then ligated together ( Fig. 1. ). The fidelity of the process is well assured.
The replication mechanism is precise and accurate because DNA polymerase also
exhibits a proofreading activity by →5 exonuclease activity, whereby it removes
any wrong nucleotide added. Another protein known as sliding clamp protein/repli-
cation factor C prevents detachment of DNA polymerase from the DNA strand but
releases it once an Okazaki fragment ends. The ends of the chromosomes have spe-
cial sequences called telomeres. The enzyme telomerase binds to the telomeres and
helps to end the replication mechanism. Finally, the primers of RNA are removed
and replaced with DNA and then ligated to the newly formed strand by DNA ligases.
When a cell's replication and repair process fails, a rare event, a permanent
change called mutation results. DNA also continuously incurs chemical changes like
insertion or deletion, substitution, depurination, deamination, dimer formation, and
so forth. The cell has several repair mechanisms in place. In the mismatch repair
system, a group of repair proteins recognizes these mistakes, cuts the damaged DNA
strand, and adds nucleotides at the  end complementary to the undamaged strand.