Composition of chromosomes

Introduction

The major components of the chromosomes are:

must read: What are Chromosomes? : Its function, and Structure

Proteins

Histones

• Histones are groups of relatively small proteins and they have a strong positive charge because they have a high content of Amino acids Lysine and Arginine. The DNA is negatively charged and the binding of the Histones with DNA is stabilized by the Ionic bonding. Histones play an important role in the structure of proteins.
• There are five main types of histones H1, H2A, H2B, H3, and H4. The mass of the histones in chromatin is approximately equal to the mass of DNA in most cells. An equal volume of H2A, H2B, H3, and H4 molecules are present in chromatin but about half that number of H1 molecules. In different kinds of Eukaryotic cells, this proportion remains constant.
• Their basic function is that they bind with the DNA and help the chromosomes maintain their shape and control the activity of genes. Particularly all the types of nuclei contain histone proteins. They have basic properties and play an important role in regulating the function of the chromosomal DNA.

Non-Histone proteins

They are primarily acidic and as regulatory molecules considered more important than the histones. In simple words, we can say that if we removed all histone portions from the chromatin the remaining portion will be known as Non-histone proteins.

There are more than a thousand types of non-histone proteins that are involved in DNA replication and gene expression. Chromatin also contains a diverse group of non-histone proteins that play a variety of structural roles for example Chromosome scaffold, enzymatic role, regulatory roles, and Chromosome replication, for example, DNA polymerase and chromosome segregation Examples are kinetochore proteins.

DNA

• DNA  which is stands for Deoxyribonucleic acid. They are the polymers of nucleotide repeat units that consist of four types of Nitrogenous bases, sugar, and phosphate. They are double stranded molecules. It has a significant role in the development and function of an organism because it contains genetic information.
• In 1953, James Watson and Francis Crick working at Cambridge University in England formulated the double helical structure of DNA. The original X-ray diffraction data was generated by Rosalind Franklin working in London at King’s college. The diffraction pattern revealed that the DNA is long, thin, and helical, and one type of structure repeated every 0.34nm. Watson and Crick used the information provided in Franklin’s picture and proposed the DNA double helical structure by building the wire model of a possible structure.
• Erwin Chargaff was interested in the Base composition of DNA. For separation and to measure the quantity of four bases he used the chromatographic methods in 1944 and 1945. The four bases of the DNA are Adenine (A), Guanine (G), Cytosine(C), and Thymine (T). He found that the Adenine number is equal to the number of Thymine (A=T) and the Guanine number is equal to the number of Cytosine (G=C). Similarly, the number of purines is equal to the number of Pyrimidines (A+G=C+T). The importance of these equivalencies is called Chargaff’s rules and these are confirmed after the discovery of DNA by Watson and Cricks.
• The defining feature of DNA is the sequence of the nucleotides along the length of the DNA.
• There are some important features of the double helical structure. It consists of a Major groove and a Minor groove and the two strands are twisting around each other. Without unfolding the double helical structure with the help of the regulatory proteins we can find the specific base sequence on DNA.
• The two strands of the DNA are antiparallel to each other. In the double strand of DNA, the 5’ carbon of one nucleotide joins to the 3’ carbon of the other nucleotide oriented in the opposite direction with the help of the Phosphodiester bond.
• The two strands of the DNA can be denatured or renatured. The two strands are bound together by relatively weak non covalent bonds so by raising the temperature and PH we can easily separate the two strands of DNA called Denaturation (Melting). The reverse process is called DNA renaturation or reannealing.
• Chromosomal DNA also contains Euchromatin and Heterochromatin.

RNA

• RNA, which stands for Ribonucleic acid, is the polymer of nucleotide repeat units consisting of one of four types of nucleotides. three are the same as in DNA but slightly differ in sugar and phosphate. Unlike DNA they are single stranded molecules. It controls the synthesis of proteins by acting as a messenger by carrying the instructions from the DNA.
• Transcription of the RNA is performed by the DNA and most of the RNA is transported to the cytoplasm and some quantity remains associated with protein and also along with the DNA. RNA is found abundantly in the cytoplasm.
• The transcription is the process by which an RNA molecule is produced called the primary transcript and before functioning in the cell undertaken many chemical changes. To explain the chemical modifications we use the term RNA processing in which we obtained the final product in the form of mature RNA product from the primary transcript. Processing follows the removal of the portions of primary transcripts and is followed by the addition of nucleotides or chemical modification of specific nucleotides. Methylation of bases or ribose group is an example of it.
• There are different types of RNA.
• The first one is mRNA which provides information for the production of proteins.
• The second one is rRNA during the process of translation instruct and initiates the assembly of polypeptides from the mRNA.
• The third one is tRNA, during the process of translation brings the correct amino acid to mRNA.
• Some RNAs are involved in RNA processing for example Small nuclear RNAs (snRNAs) and Small nucleolar RNAs (snoRNAs).
• Some RNAs are involved in the regulation of gene expression for example MicroRNAs (miRNAs) and Small interfering RNAs (siRNAs).

Difference between DNA and RNA

DNA	RNA
Nitrogenous bases are Adenine, Thymine, Cytosine, and Guanine	Nitrogenous bases are Adenine, Uracil, Cytosine, and Guanine
It is double stranded	It is single stranded
De oxy ribose sugar is present	Ribose sugar is present
On the basis of the arrangement of bases, there is only one type of DNA with numerous variations	Three types are present (mRNA, tRNA, and rRNA)
Contains genetic information	Responsible for protein synthesis
Made up of millions of nucleotides, larger in size	Made up of hundreds of nucleotides, smaller in size
In each cell of a species, its amount or quantity remains constant	In different cells, quantity is also different
It is present in the nucleus, chloroplast, and in mitochondria.	It is present in the nucleus, cytosol, ribosomes, mitochondria, and nucleolus
Life span is high	Life span is less

The molecular composition of the chromosome is based on Folded Fiber Model and this model is proposed by Du Praw in 1965 and is a widely accepted model. Chromosomal DNA is coiled hierarchically. structure and composition of Chromosome is highly ordered in the Eukaryotic cell and each chromosome undergoes a certain level of condensation or complexity.

DNA is complex with several proteins and to achieve this level of complexity DNA undergoes coiling and sub coiling to form the Chromatin. DNA is a 2nm thick double helix. During the Interphase when the DNA is in highly extended form, the double helix DNA undergoes at least two levels of coiling followed by the binding of specific histone proteins. The first step is the formation of a 10nm thick filament and then this thick filament is coiled into 30nm thick chromatin fiber. The chromatin fiber undergoes the looping and the Scaffold of non-histone proteins provides them support.

Basic Unit of Chromatin structure

Nucleosomes

• The nucleosome is made up of eight Histone molecules or proteins (two each of Histone H2A, H2B, H3, and H4) and these histone proteins are wrapped by a stretch of 147 base pairs of double stranded DNA. The DNA and protein molecules bind with each other by unstable ionic nature and this bonding or linkage is known as Salt linkage. For example, Ca++ and Mg++ are specific metallic ions.
•  Nucleosomes bind with other nucleosomes with the help of Linker DNA having a short length of 8 to 147 base pairs. The length of the Linker DNA is varied according to the different species. The diameter of the Nucleosome is 10nm. Each histone protein in the nucleosome has tails known as N-terminal tails. Histone molecule tails are made up of specific amino acids and can undergo various types of translational modifications preferably Acetylation, Phosphorylation, and Methylation with the help of these N-terminal tails. Different proteins can bind to chromatin and as a result, affects the chromatin condensation and transcriptional activity.
• Nucleosomes are packed together to form chromatin fibers and chromosomes.
• The first step in the Nuclear DNA packaging is the formation of the nucleosomes. Chromatin fibers are just like Beads on a string and are 10nm in diameter but the chromatin of the intact cell forms a thick fiber that is 30nm in diameter. The 10nm and 30nm forms of the chromatin fiber can be interchanged by changing the solution’s salt concentration in the preparation of isolated chromatin. If we remove the H1 histone molecule we cannot prepare the 30nm fiber which tells us that for the packing of the nucleosome into 30nm fiber we need the H1 histone.
• An irregular, three dimensional zigzag structure that has the ability to combine with the surrounding fibers is formed when the 30nm fibers are packed together. The folding of the 30nm fibers results in the formation of DNA loops which consist of 50,000 to 100,000 base pairs in length.
• Cohesin is a protein that plays an important role in maintaining these DNA loops and performing another function prior to anaphase in dividing cells keeping the chromosomes attached to each other. The long loops of the DNA are attached with a chromosomal scaffold which is formed by the periodic binding of the DNA with non- histone insoluble network necessary for maintaining the specific arrangement of the loop.
•  With the help of an electron micrograph of chromosomes, we can easily visualize the loops which are separated from the dividing cells and from which all histone and non-histone proteins are removed. Polytene chromosomes a specialized type of chromosome having thousands of DNA strands can also be used for the visualization of the loop and these chromosomes are not associated with cell division. The active DNA is less tightly packed than the inactive DNA because the proteins involved in gene transcription can easily access the active DNA.

Folded Fiber Model

Folded Fiber model is proposed by Du Praw in 1965. Within a nucleus, DNA is present and the length is about one meter or more if we extend it completely while the diameter of the nucleus is not more than 5-10nm. The folding of this wide length of DNA into a nucleus that is a million times smaller than the DNA was a very odd thing. Each chromatin fiber has an average diameter of 230Å and consists of a DNA molecule.

Chromatin fibers are present inside the chromatin. Two sister chromatin fibers are produced during the interphase when the DNA of the chromatin fiber replicates. Sister chromatin fibers remain joined to each other in the Centromeric region but when replication occurs in the centromeric region the sister chromatid fibers are separated in this way.

Two sister chromatids are formed from two sister chromatids during cell division when extensive folding is undertaken by the two sister chromatids in an irregular manner. The sustainability and thickness of the chromatin fiber are increased by folding the chromatin fibers because the folding reduces their length. This folded structure further undergoes supercoiling and as a result, further reduces the length and thickness of the chromosomes increases.

1. In the late 1960s, fraction studies were carried out by Maurice Wilkins on the folding process and it reveals that the purified chromatin fibers do not consist of only DNA and Histones. Wilkins concluded that on the DNA, histones impose a repeating structural organization.
2. In 1974, Ada Olins and Donald Olins isolated the chromatin fibers and publish micrographs of chromatin fibers. They isolated the chromatin fibers in such a way that they prevent the use of harsh solvents that were used in early times for the preparation of the chromatin for the microscopic examination. They observe the small particles bind with each other with help of the filaments and this was actually the appearance of the chromatin fibers. This appearance was given the name “Beads on String” suggesting that the beads represent the Histone proteins and thin filaments represent the DNA wrap around the histone core. So we can say that Nucleosome is the basic structural unit of chromatin. It is made up of a coil of DNA wrapped around a histone core.
3. Now the next question was raised that the nucleosomes were the normal component of the chromatin or any impurity during the process of sample preparation. Dean Hewish and Leigh Burgoyne were the two scientists who performed the experiment on Rat liver. They found that the rat liver contains an enzyme that is capable of cleaving the DNA into chromatin fibers. These scientists exposed chromatin to this nuclease enzyme. They purified the partially digested DNA and remove the chromatin proteins. The purified DNA is then examined by gel electrophoresis.

Conclusions

They found that each fragment of the DNA has a distinct pattern. The smallest piece of the DNA is 200 base pairs long and the remaining fragments are exact multiples of 200 base pairs. They conclude that the fragment pattern is not generated by the protein free DNA. Along the DNA molecule, chromatin proteins are wrapped in a regular pattern and this pattern is repeated after every 200 base pairs. The DNA which is present between the histone proteins is digested by the nuclease and fragments are produced which are 200 base pairs in length.

Reference

Hardin, J., Bertoni, G., & Becker, W. M. (2018). Becker’s world of the cell. Pearson.