Dna Polymerase I

Lagging strand template strand is paired with the 5' end of the other. One newly replicating strand (the leading strand) is pointing in the "right" direction to grow continuously at its 3' end as the fork opens up. But the other strand (the lagging strand) is pointing in the "wrong" direction: As the fork opens up further, its exposed 3' end gets farther and farther away from the fork, and an unreplicated gap is formed, which would get bigger and bigger if there were not a special mechanism to overcome this problem.

Single-strand DNA-binding proteins make the templates available to primase and DNA polymerase III.

11.15 Many Proteins Collaborate at the Replication Fork Several proteins in addition to DNA polymerase III are involved in DNA replication. The two molecules of DNA polymerase (red) are actually part of the same complex.

The lagging strand is synthesized from Okazaki fragments

Synthesis of the lagging strand requires working in relatively small, discontinuous stretches (100 to 200 nucleotides at a time in eukary-

Lagging RNA primer strand

Primase forms an RNA primer. j

Primase ^ RNA primer

Primase forms an RNA primer. j

Primase ^ RNA primer

Rna Primer And Primase
11.17 The Lagging Strand Story In bacteria, DNA polymerase I and DNA ligase cooperate with DNA polymerase III to complete the complex task of synthesizing the lagging strand.

otes; 1,000 to 2,000 at a time in prokaryotes). These discontinuous stretches are synthesized just as the leading strand is, by the addition of new nucleotides one at a time to the 3' end of the new strand, but the synthesis of this new strand moves in the direction opposite to that in which the replication fork is moving. These stretches of new DNA for the lagging strand are called Okazaki fragments, after their discoverer, the Japanese biochemist Reiji Okazaki. While the leading strand grows continuously "forward," the lagging strand grows in shorter, "backward" stretches with gaps between them.

A single primer suffices for synthesis of the leading strand, but each Okazaki fragment requires its own primer. In bacteria, DNA polymerase III synthesizes Okazaki fragments by adding nucleotides to a primer until it reaches the primer of the previous fragment. At this point, DNA polymerase I (the one discovered by Kornberg) removes the old primer and replaces it with DNA. Left behind is a tiny nickā€”the final phosphodiester linkage between the adjacent Okazaki fragments is missing. The enzyme DNA ligase catalyzes the formation of that bond, linking the fragments and making the lagging strand whole (Figure 11.17).

Working together, DNA helicase, the two DNA poly-merases, primase, DNA ligase, and the other proteins of the replication complex do the job of DNA synthesis with a speed and accuracy that are almost unimaginable. In E. coli, the replication complex makes new DNA at a rate in excess of 1,000 base pairs per second, committing errors in fewer than one base in 106, or one in a million.

Telomeres are not fully replicated

As we have just seen, replication of the lagging strand occurs by the addition of Okazaki fragments to RNA primers. Beyond the very end of a linear DNA molecule, however, there is no place for a primer to bind (i.e., there is no complementary DNA strand). So the new chromosome formed after DNA replication has a bit of single-stranded DNA at each end (Figure 11.18a). This situation activates mechanisms that cut off the single-stranded region, along with some of the intact double-stranded end. Thus, the chromosome becomes slightly shorter with each cell division.

In many eukaryotes, there are repetitive sequences at the ends of chromosomes called telomeres. In humans, the telomere sequence is TTAGGG, and it is repeated about 2,500 times. These repeats bind special proteins that maintain the stability of chromosome ends. Each human chromosome can lose 50-200 base pairs of telomeric DNA after each round of DNA replication and cell division. After 20-30 divisions, the chromosomes are unable to take part in cell division, and the cell dies. This phenomenon explains in part why cells do not last the entire lifetime of the organism: Their telomeres shorten.

Yet constantly dividing cells, such as bone marrow and germ line cells, maintain their telomeric DNA. An enzyme, appropriately called telomerase, catalyzes the addition of any lost telomeric sequences (Figure 11.18b). Telomerase contains an RNA sequence that acts as a template for the telo-meric repeat sequence.

Telomerase is expressed in more than 90 percent of human cancers and may be an important factor in the ability of can-

Telomerase

/ RNA primer

Telomeres And Telomerase

Telomerase

/ RNA primer

Removal of the RNA primer leads to the shortening of the chromosome after each round of replication. Chromosome shortening eventually leads to cell death.

11.18 Telomeres and Telomerase (a) Removal of RNA primer at the 3' end of the lagging strand leaves a region of DNA unreplicated.

(b) The enzyme telomerase binds to the 3' end and extends the lagging strand of DNA. An RNA sequence embedded in telomerase provides a template so that, overall, the DNA does not get shorter.

(c) Bright fluorescent staining marks the telomeric regions on these blue-stained human chromosomes.

Removal of the RNA primer leads to the shortening of the chromosome after each round of replication. Chromosome shortening eventually leads to cell death.

Unreplicated Strand

11.18 Telomeres and Telomerase (a) Removal of RNA primer at the 3' end of the lagging strand leaves a region of DNA unreplicated.

(b) The enzyme telomerase binds to the 3' end and extends the lagging strand of DNA. An RNA sequence embedded in telomerase provides a template so that, overall, the DNA does not get shorter.

(c) Bright fluorescent staining marks the telomeric regions on these blue-stained human chromosomes.

cer cells to divide continuously. Since most normal cells do not have this ability, telomerase is an attractive target for drugs designed to attack tumors specifically.

There is also interest in telomerase and aging. When a gene expressing high levels of telomerase is added to human cells in culture, their telomeres do not shorten. Instead of dying after 20-30 cell generations, the cells become immortal. It remains to be seen how this finding relates to the aging of a large organism.

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  • HENRIIKKA
    Why do ends of chromosomes get shorter after each round of DNA replication?
    7 years ago

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