Introduction to Molecular Biology: From Cells to Atoms - Anthony Rees, Michael Sternberg 2002

Nucleic Acids and Genes
Packaging of Genetic Material

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Fig. 26.1.

Prokaryotic Cells contain a single copy of their genomic DNA and are thus haploid. The DNA of a Prokaryotic Cell, which encodes all cellular Proteins and Nucleic Acids (rRNA, tRNA, etc.), is part of the chromosome. For instance, E. coli contains a single chromosome consisting of a complex of DNA, RNA, and proteins. The DNA is a circular molecule containing 4.6 ∙ 10⁶ nucleotide pairs (or 4600 kilobase pairs, kbp). While the diameter of such a circle would be approximately 1 mm, E. coli itself has a transverse diameter of less than 2 µm; consequently, its DNA must be tightly packed (condensed) to fit inside The Cell.

Eukaryotic cells are more complex than prokaryotic cells and, with the exception of Gametes, all contain two identical copies of The Genome; in other words, they are diploid. A nematode cell contains approximately 40 times, and a salamander cell about 40,000 times, more DNA than E. coli. A human cell contains roughly 700 times more DNA than E. coli. Because humans are generally considered to be more complex organisms than salamanders, we must conclude that total cellular DNA content serves as a very rough indicator of an Organism's biological complexity.

In eukaryotic cells, DNA is located in The Nucleus as a set of separate fragments called Chromosomes. Each chromosome can contain anywhere from 400 (in Yeast) to 100,000 kbp (in humans) of DNA. If all the cellular DNA in a simple double-helix conformation were stretched out into a single line, it would be excessively long (1.74 m for a human cell) and could not possibly fit within the nucleus; therefore, chromosomes must exist as highly condensed structures.

Viruses can contain either single-stranded or double-stranded DNA, which may be continuous or fragmented. In some viruses (e.g., TMV, reoviruses; Chapter 5), the genetic material is RNA, which can also be either single-stranded or double-stranded.

The number of genes in various organisms can be estimated based on the average Gene length of 1 kbp. Table 26.1 provides the corresponding figures.

The Condensation of double-stranded DNA, which reduces the longitudinal dimensions of the molecule by a factor of 10,000, is achieved through one of two mechanisms: spheroidal winding (i.e., coiling along a spheroidal generator) or The formation of supercoiled DNA. Spheroidal winding occurs essentially only in viruses, whereas supercoiling takes place in all prokaryotic and eukaryotic cells, as well as in many viruses that infect eukaryotes.

Table 26.1

Spheroidal DNA winding results in the formation of a compact "spool". In bacteriophage T4, the DNA is coiled into a double-layered spool inside the icosahedral phage HEAD. First, the turns of the outer layer are formed, followed by the inner layer, leaving a certain clearance between the layers. The remaining end of the DNA passes through both layers and exits into the viral tail tube via a central aperture (Chapter 4). In other viruses, DNA may coil somewhat differently than in T4, but in a very similar manner overall.

Supercoiled (superhelical) DNA is formed when extra turns (supercoils) are introduced into The Double Helix. This induces stress within the molecule, which manifests notably in the twisting of the double-helix axis itself into a secondary helix (supercoiling), as illustrated in Fig. 26.2 for a covalently closed circular double-stranded DNA molecule.

Fig. 26.2.

Stabilization of compact DNA Conformations is essential for both spheroidal winding and supercoiled DNA, because such tight packing brings negatively charged phosphate groups into close proximity. The mutual electrostatic repulsion between these groups is counteracted by their binding to positively charged proteins and small molecules known as Polyamines. Polyamines are found in all prokaryotic and eukaryotic cells without exception, though they are absent in many viruses. Examples of polyamines discovered in eukaryotic cells include spermidine, The Structure of which is shown in Fig. 26.3. Specific proteins that stabilize the DNA molecule are present in all cell types, but in only a few viruses (e.g., SV40 virus).

¹ According to recent data, the actual number of human genes is significantly lower. — Transl. note.

Fig. 26.3.

DNA packaging in prokaryotic cells is likely universal in character; let us examine how it occurs in the bacterium E. coli, for instance. The supercoiled DNA segment, complexed with histone-like (see below) DNA-binding proteins and polyamines, forms a "bead" approximately 12 nm in diameter. Within the bead, the axis of the DNA double helix is twisted to form roughly six turns. These beads are interconnected by unsupercoiled stretches of DNA, thereby creating a structure reminiscent of a necklace, rosary, or beads on a string. This bead-organized DNA then forms large loops, which are stabilized and ultimately condensed through interactions with specific proteins and RNA.

DNA packaging in eukaryotic cells involves the association of supercoiled DNA with various proteins (Histones) to form a complex called Chromatin. Chromatin, in turn, forms a solenoid-like structure that associates with chromosomal structural proteins, culminating in the complex known as a chromosome.

Chromatin consists of double-stranded DNA wrapped around specialized core particles made of specific proteins called histones, forming a bead-like structure referred to as a chromatin fiber. Each bead, known as a nucleosome, is about 10 nm in diameter (e.g., in E. coli). This condensation of DNA into chromatin reduces its longitudinal dimensions by approximately 5-6 fold.

A nucleosome is a segment of double-stranded DNA approximately 200 Base Pairs long wrapped around a protein core consisting of eight histone molecules. Its structure, first elucidated by Klug and co-workers, is depicted in Fig. 26.1. Nucleosomes have been found in all eukaryotic cells and in certain viruses that infect eukaryotes. Histones are basic proteins with molecular weights ($M_r$) ranging from 11,300 to 21,000. There are five known types of histones: H1, H2A, H2B, H3, and H4. The nucleosome core contains two molecules each of histones H2A, H2B, H3, and H4. The outward-facing surfaces of these protein molecules carry positive charges, forming a stabilizing scaffold around which the negatively charged DNA molecule can wrap. Histone H1 is located at the DNA segments connecting adjacent nucleosomes; these segments are therefore called linker DNA. Because H1 does not appear to be involved in stabilizing the core Chromatin Structure, it likely plays a role—along with certain non-histone proteins—in regulating the transcriptional activity of chromatin.

The solenoid-like structure of cellular chromatin was proposed to explain the Water/144.html">Origin of the so-called 20–30 nm heterochromatin fibers observed under an Electron microscope in interphase nuclei (Chapter 29). Each such solenoid is formed by the further coiling of the DNA double helix, which is already supercoiled at the nucleosome level. There are approximately 6 nucleosomes per turn of the solenoid. This condensation step further reduces the longitudinal dimensions of the DNA by about 40-fold compared to naked double-stranded DNA. Currently, heterochromatin is considered to be a transcriptionally inactive form of chromatin. Because the length of uncoiled double-stranded DNA and that of the same DNA in a highly condensed chromosome differ by a factor of 5,000 to 10,000, we still need to account for the remaining 100–200 multiplier [40 ∙ 100(200) = 5,000(10,000)]. However, the exact mechanism by which the solenoid-like chromatin structure undergoes further condensation into its chromosomal form remains unknown.

Chromosomal structural proteins form a scaffold upon which the final condensation of chromatin occurs, producing the tightly packed structure characteristic of eukaryotic chromosomes. This proteinaceous Skeleton remains intact even after all histones are removed.