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Essay On Dna Replication And Structure

DNA and Replication

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DNA and Replication

You pose an interesting question – There are different types of Human
DNA – which there are various classifications, Chromosomal DNA and
Mitochondrial DNA. There is also the DNA present from normal flora
microorganisms such as bacteria, viruses, mites, etc. Some of this
microorganism DNA may be significant, such as E. coli DNA in the gut
or Staphylococcus DNA on the skin. You even have DNA present from
viruses of bacteria such as phage DNA. Some human viruses may be
present in blood cells such as EBV, CMV in nerve cells like herpes
simplex 1, in skin cell like HPV (human papilloma virus) or integrated
into the Human Chromosomal DNA such as various retroviruses, like
human foamy virus, HTLV or HIV

Within Chromosomal DNA there is DNA that codes for genes- exons (mRNA
coding) and non coding regions called introns. There are regions of
DNA within the introns that are called endogenous retroviruses – these
regions have great similarity to retroviruses and may have disease
implications.

2

An Okazaki fragment is a relatively short fragment of DNA that is
created by primase and Pol III along the lagging strand (see DNA
replication). They are later removed by RNAse H, and the last
ribonucleotide is removed by and synthesized by Pol I. The nick, or a
broken phosphodiester bond remaining between the fragments is linked
together by DNA ligase

The replication fork is a structure which forms when DNA is ready to
replicate itself. It is created by topoisomerase, which breaks the
hydrogen bonds holding the two DNA strands together. The resulting
structure has two branching "prongs", each one made up of a single
strand of DNA. DNA polymerase then goes to work on creating new
partners for the two strands by adding nucleotides.

A primer is a nucleic acid strand (or related molecule) that serves as
a starting point for DNA replication

Oka-what? Another Look at Okazaki Fragments.

As you have already learned, the two strands of DNA are antiparallel.

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This presents real difficulties during replication. The DNA polymerase
III enzyme synthesizes most of the DNA. The enzyme has two subunits
because both strands of parental DNA must be replicated in the same
place at the same time.

Because DNA can only be synthesized in the 5' to 3' direction, one of
the strands (the 3' to 5' strand) can be copied continuously and is
called the leading strand, while the other, the lagging strand, is
synthesized in fragments so that the 5' to 3' polymerization leads to
overall growth in the 3' to 5' direction. This may be accomplished by
a looping of the template for the lagging strand (see Figure below).
The lagging strand would then pass through the polymerase site in the
same direction as the leading-strand template in the other subunit.
DNA polymerase III would then have to let go of the lagging strand
template (after about 1,000 nucleotides have been added to the lagging
strand). A new loop would then be formed. The gaps between fragments
of the lagging strand are filled by another polymerase, DNA polymerase
I, and the enzyme DNA ligase joins the fragments.

The looping of the template for the lagging strand enables the DNA
polymerase III enzyme (colored yellow) at the replication fork to
synthesize both daughter strands in the 5' to 3' direction. The
leading strand is shown in blue, the lagging strand in green.

[IMAGE]

[IMAGE]

[IMAGE]This figure (at left, click to enlarge) may help explain why
the lagging strand must be looped during DNA synthesis. A key point is
that DNA polymerase III in the replication fork is organized into a
dimer - a pair of molecules linked together. Both DNA polymerases
(pink structures in the diagram) are oriented in the same direction
and, as you know, will move along the parent strand from the 3' end to
the 5' end, synthesizing new DNA in the 5' to 3' direction. The purple
ring in the diagram is DnaB helicase, an enzyme that is linked to the
DNA polymerase dimer and functions in separating (unwinding) the two
parent strands from each other. As helicase moves (relative to the
DNA) along, it opens up the replication fork, allowing the polymerases
to access the template strands. Since the polymerases are reading the
templates from right to left in the diagram, the only way for the
bottom template strand to pass through the polymerase in the required
3' to 5' direction is to loop the DNA strand as shown.Upon completion
of one Okazaki fragment the polymerase release that fragment and the
DNA strand is pulled forward bringing the next primed region (green)
into contact with the polymerase and allowing synthesis of a new
fragment to begin.

3

Frameshift mutation: A deletion or insertion of any number of bases
other than a multiple of three bases has a much more profound effect.
Such frameshift mutation result in a complete change in the amino acid
sequence downstream from the point of mutation, instead of simply a
change in the number of amino acids. (Figure 1)

Normal

THE BIG RED DOG RAN OUT.

Missense

THE BIG RAD DOG RAN OUT.

Nonsense

THE BIG RED.

Frameshift -- deletion

THE BRE DDO GRA.

Frameshift -- insertion

THE BIG RED ZDO GRA

Causes of mutations: Mutations are caused by substances that disrupt
the chemical structure of DNA or the sequence of its bases.
Radiation, various chemicals, and chromosome rearrangements are some
of the many sources of mutation.

Mutation rates: All of us are subjected to mutagenic events
throughout our lifetime. Depending upon the type of mutation, the
frequency ranges from 10-2/cell division to 10-10/cell division. Our
cells have numerous mechanisms to repair and/or prevent the
propagation of these mutations.

http://www.uic.edu/nursing/genetics/Lecture/Molecular/mutation.htm

Another type of mutation involves either the insertion or deletion of
one or more (some number that is not a multiple of three) nucleotides
into a DNA sequence. This type of mutation is known as a frameshift
mutation. For an illustration of how devastating this type of
mutation can be if it occurs in the coding region of a gene, delete
the w from the sentence below.

The cow jumped over the moon.
becomes
The coj umpedo vert hem oon.

The insertion of nucleotides in multiples of three, if not corrected
during the culling of introns from messenger RNA, will cause the
insertion of an extra amino acid for each three additional
nucleotides. Trinucleotide repeats are a sequence of three
nucleotides that repeat in tandem and vary in the the number of
repeats. Trinucleotide repeat mutations are known to cause at least
eight genetic disorders affecting the nervous or neuromuscular
system. For more about this topic see
http://prl.humc.edu/obgyn/public/genetics/trirep.htm .

YAP an alu insertion

http://www.contexo.info/DNA_Basics/Mutations.htm

Frameshift mutations

* If DNA is copied incorrectly by the mRNA and a base is deleted (or
added) it causes all other bases to shift over one and code for
incorrect amino acid sequences after the error.

* Frameshift mutation- mutation in which a single base is added or
deleted from DNA

* Frameshift mutations much worse than point mutation due to
quantity of amino acids affected

http://www.northstar.k12.ak.us/schools/wvh/science/dcox/Chapter%20outlines/bioch11.htm

* Frameshift Mutations: Additions or deletions of one or more
nucleotides.

* May result in "garbage" genes, as the entire amino acid sequence
in the code after the change is devastated.

* Large deletions may remove a single amino acid, or an entire
chunk of chromosome. The most common mutation that causes severe
cystic fibrosis deletes only a single codon.

* Real examples of missense, nonsense, and frameshift mutations:

Hemoglobin mutants and Hemoglobin molecule

A note of caution. These examples show the NON-template DNA sequence
rather than the template DNA sequence as in our previous examples.
This is a standard used by DNA scientists. To get the mRNA codons,
just change the Ts to Us.

* Some genes have repeated base sequences, and the number of these
may increase each generation. These expanding genes are
responsible for increasingly severe cases of muscular dystrophy
(CTG repeats), Huntington disease (CAG repeats), and Fragile X
syndrome (CGG repeats).

Fragile X Syndrome:
6-50 CGG repeats in an unaffected individual
50-200 CGG repeats in a carrier
>200 CGG repeats in an affected individual

http://www.life.uiuc.edu/bio100/lectures/s04lects/15s04-mutations.html



4Mitosis vs Meiosis
===================


Mitosis


Mieosis


Produces body cells(Somatic cells)

Produces sex cells(Gametes)


Daughter cells diploid(2N)

Daughter cells haploid(N)


Two daughter cells produced

Four daughter cells produced


In metaphase chromosomes line up singley


In metaphase I chromosomes line up as homologous pairs(synapsis)
The two double chromosomes are called a tetrad when they are lined up
side-by-side.
Crossing over occurs during the formation of the tetrad


One nuclear division

Two nuclear divisions


Produces cells for growth and repair

Produces cells for sexual reproduction


Daughter cells have two sets of chromosomes(pairs)

Daughter cells have only one member of each pair of chromosomes


Daughter cells are genetically identical to the parent cell

Daughter cells have one-half of the genes from the parent cell


Insures that all daughter cells are genetically identical

Generates genetic diversity through crossing over and ramdom
seperation of homologous pairs of chromosomes



DISCUSSION QUESTIONS FOR CHAPTERS 1, 2, 3, 4, 5, 6, 7, 8

1: SOME PRINCIPLES

1. How is it possible to discover the functions of the 'non-coding' sequences in and around a gene? To what extent have these techniques yielded satisfactory answers?

2. Repetitive DNA sequences are a major component of mammalian genomes. Describe the different classes of such sequences, and outline what - if any - biological function they may serve.

3. Write an essay on the recognition of information in nucleic acids.

4. What forces maintain the structure of a DNA duplex?

5. Illustrate how differences between the structure of DNA and RNA are reflected in the ways that proteins interact with them.

6. Genes have been defined in many different ways over the years. Describe as many of these ways as you can. What definition is appropriate today?

7. What is DNA supercoiling? How is it generated? What are its biological roles?

8. Discuss redundancy in the genome, and the roles that it plays.

9. Estimates for gene numbers suggest that mammals have four times more genes than flies, and ten times more than yeast. Discuss.

10. What is the role of the nuclear membrane?

11. What are the three primary lineages of the living world, and how do they differ?

12. What design principles are used in the construction of large biological structures like virus particles, the cytoskeleton, and chromosomes.

2: STRUCTURE

1. How is the structure of the nuclear pore related to its function?

2. Discuss how proteins are imported into the nucleus.

3. Describe what we know about the synthesis and processing of ribosomal RNA.

4. Would you describe the nucleolus as a ribosome factory, when we know so little about the assembly of ribosomal RNA into a ribosome?

5. Describe the hierarchies of organization of DNA from the double helix to the chromosome. What problems does the organization pose for transcription and replication?

6. How is the structure of DNA in the isolated 'nucleoid' related to that found in vivo?

7. What is the evidence that clusters of chromatin loops are organized into 'clouds' around transcription 'factories'?

8. The cytoplasm contains a well-characterized skeleton. Discuss the evidence for and against the existence of an analogous skeleton within the nucleus.

9. Write an essay on compartmentalization in the nucleus.

10. Why has it been so difficult to determine the structure of a eukaryotic chromosome, whether in interphase or mitosis?

11. Most eukaryotic chromosomes have similar shapes, even though they may contain very different amounts of DNA. How adequately do current models for the organization of the DNA fiber within a chromosome account for its general shape?

12. Discuss current models for the structure of chromatin and chromosomes. How far do they account for the various functions of DNA?

13. What are polytene chromosomes, and how are they formed?

14. Why do mitotic chromosomes have the shape they do?

15. Discuss telomeres in terms of their discovery, location, universality, duplication, and relationship with ageing and cancer.
 

3: REPLICATION

1. Discuss the evidence for and against the idea that active DNA polymerases are organized into factories.

2. What problems does the double-helical structure of DNA pose for the process of replication?

3. Describe the roles of the different proteins involved in replicating a DNA duplex.

4. How does the process of replication on one side of a replication fork differ from that on the other?

5. DNA polymerases make mistakes. Describe the mechanisms that ensure that parental and daughter duplexes have the same DNA sequences.

6. Describe how the origins of replication in pro- and eu-karyotes can be defined.

7. Compare and contrast the origins of replication found in simple organisms with those of mammalian cells.

8. 'There is no such thing as a specific origin of DNA replication in eukaryotes'. Discuss.

9. Discuss the role played by transcription during replication.

10. Discuss the problems associated with replicating the ends of a chromosome. How are these problems solved?
 

4: TRANSCRIPTION

1. Describe the topological problems associated with transcribing a double-helical template. How are these problems solved?

2. Outline the molecular events that lead to the synthesis of a primary transcript by RNA polymerase II, and describe how evidence for the process was obtained.

3. Discuss the evidence for and against the idea that active RNA polymerases are organized into factories.

4. Describe the properties of the three eukaryotic RNA polymerases and their templates.

5. Comparison of the promoter sequences of a family of mammalian genes reveals that all share a sequence of eight nucleotides. Outline how you would test experimentally the possible role of this octamer sequence in regulating the expression of these genes.

6. Outline the modifications that occur to ribosomal RNA as it matures. How were these modifications discovered?

7. The initiation of transcription by eukaryotic RNA polymerases requires the assembly of a large complex. Outline the order of events that result in initiation, and indicate the type of molecular interactions that are involved.

8. RNA polymerases make mistakes. Describe the mechanisms that ensure that messages contain the correct coding information.

9. To what extent can a transcriptional activity found in vivo be reproduced in vitro?

10. Discuss the role played by the C-terminal domain of RNA polymerase II in the production of a transcript.

11. Describe how a transcript made by RNA polymerase II is modified.

12. How are primary transcripts processed and what roles do such modifications play?

13. Describe the role played by RNA:RNA interactions in the removal of introns from the primary transcript of eukaryotic genes transcribed by RNA polymerase II.
 

5: REPAIR

1. Describe the lesions that are commonly found in DNA. What are the consequences if they go unrepaired?

2. Discuss the advantages and disadvantages of the different approaches that have been used to detect the ways in which damaged templates are normally repaired.

3. Illustrate how the study of human disease has helped us to understand the different pathways involved in repairing damage in DNA.

4. Compare and contrast the major pathways involved in repairing damage in human DNA.

5. What are the consequences of a failure to repair damaged templates?

6. Genomes seem to contain more genes involved in repairing DNA than in replicating it. Why?

7. Outline the evidence that some repair of damage in DNA is coupled to transcription.
 

6: REGULATION OF GENE EXPRESSION

1. The expression of bacterial genes is controlled by the action of diffusible repressors and activators. To what extent is the expression of mammalian genes controlled similarly?

2. How true is the statement that all cells in a mammal contain the same genetic information?

3. Outline the various levels at which the expression of genes is controlled? What methods would you use to identify which control mechanisms were operating in a particular case?

4. The differentiated state is generally stable and can be inherited from one somatic cell to another. What mechanisms might account for this stability and how might you distinguish experimentally between them?

5. Describe the experimental approaches that have been used to analyze how gene expression is regulated at the level of the nucleosome (and/or) chromatin loop?

6. How do covalent modifications of histones and DNA affect gene expression?

7. How far has a detailed knowledge of the nucleotide sequence in and around genes helped to explain their tissue-specific expression?

8. Discuss the relative importance of cis- and trans-acting factors in the control of transcription.

9. Discuss the advantages and disadvantages of the various approaches being used to obtain an understanding of tissue-specific gene expression?

10. Are locus control regions any different from transcriptional enhancers?

11. Describe the experimental approaches used to define enhancers and locus control regions, and explain how the functions of the two sequences differ.

12. What are the major factors underlying the inactivity of heterochromatin?

13. 'Methylation of DNA results from, but does not cause, differentiation.' Discuss.

14. Assess the significance of DNA methylation as a mechanism for suppressing gene expression.

15. How close are we to a complete molecular definition of the inactivity of heterochromatin?

16. Outline the various mechanisms that are involved in creating (and/or maintaining) the differentiated state.

17. 'The techniques of structural biology have told us little about the regulation of gene expression that we did not already know'. Discuss. 

18. What does the birth of the first parthenogenetic mouse tell us about imprinting and mammalian development?

19. Cellular protein levels can be controlled by regulating the rate of translation.  Give some examples of the mechanisms involved, and discuss the experimental approaches used to confirm that control is exerted at the level of translation.

20. To what extent does the position of a gene in the genome determine gene expression?   Outline the experimental approaches that have been used to answer this question.

21. An enduring idea in biology sees genomes as looped, with the ties that maintain loops as barriers between different functional domains.  What is the evidence for looping, and how might those barriers work?

22. A histone 'code' is thought to regulate gene expression. Describe the experimental approaches that have been used to establish how this code might operate.

7: THE CELL CYCLE

1. Discuss the role that microtubules play in chromosome segregation.

2. The spindle contains millions of moving parts. How are these movements controlled?

3. Centromeres exhibit a bewildering structural variation. What are their main functions?

4. The cell cycle is regulated by the reversible phosphorylation of proteins. Discuss.

5. How is the synthesis of DNA controlled in eukaryotes?

6. Review the evidence supporting current models for the initiation of DNA replication in eukaryotic cells.

7. Compare the checkpoints in the cell cycles of yeast and man.

8. Assess the evidence that the mechanisms for controlling passage through the cell cycle are conserved in eukaryotes.

9. Review the mechanisms that ensure orderly progression through the cell cycle.

10 .Discuss the evidence that genetic defects are responsible for malignancy.

11. Describe the advantages and disadvantages of the various approaches being used to identify genes involved in cancer.

12. Discuss the view that malignancy results from an imbalance in the activity of oncogenes and anti-oncogenes.

13. Cancer is a multi-step process. Discuss.

14. How have studies of the nematode, Caenorhabditis elegans, contributed to our understanding of apoptosis? How does the process in the worm differ from that in higher vertebrates?

15. How is the apoptotic machinery controlled?

16. 'Cancer chemotherapy owes nothing to molecular biology.' Discuss.

17. How has the study of developmental biology impinged upon our understanding of cancer?


8: MEIOSIS AND RECOMBINATION

1. Compare and contrast the processes of mitosis and meiosis.

2. Discuss the roles that the synaptonemal complex and the chiasma play during meiosis.

3. Describe the mechanisms involved in the exchange of genetic information from one chromosome to another.

4. Describe the phenomenon of gene conversion in yeast.

5. How effectively do current models account for the properties of meiotic and mitotic recombination?

6. The breaking and joining of DNA are widespread in both prokaryotes and eukaryotes. What do we know of the various mechanisms that are used in these processes?

7. How do chromosomes pair?

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