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138 Cards in this Set

  • Front
  • Back
How are the biological roles of DNA/RNA different from other classes of biological molecules like proteins and carbohydrates?
DNA and RNA's function is the storage and transfer of information and the regulation of biochemical processes
Osborn Avery experiment
Treatment of R-form of penumococcus bacteria with pure DNA extracted from the S-form results in its transformation into the S-form that was inheritable.

Demonstrated that DNA was the transforming agent as well as the material responsible for transmitting genetic info from one generation to the next.
Consists of a pyrimidine ring fused to an imidazole ring

Includes adenine and guanine
Only consists of a six-membered nitrogen-containing ring

Includes Cytosine, Uracil (RNA), Thymine (DNA)
What is the major form of DNA bases found?
Major form of DNA bases are in amino-keto form
What is the minor form of DNA bases found?
Tautomeric form due to rearrangement of the double bonds
Tautomeric form ratio
A small fraction of each base exists in its tautomeric form
Deoxyribose + DNA base
Ribse + DNA base
Difference between Ribose and Deoxyribose sugars
At the 2` position, the deoxyribose does not have a hydroxyl group.

Ribose is less stable because the 2` hydroxyl group is attacked, leading to degradation.

The deoxyribose has enhanced stability of ring structure because of 2` deoxy
What is the linkage that is between the nucleotide base and the sugar?
N-beta-Glycosidic bond joins the Nitrogen of the base to C1` on the sugar
What are the phosphate of nucleosides derivatives of?
Phosphoric acid - H3PO4
What is the linkage that is between two different nucleosides?
Phosphodiester bonds
DNA base + deoxyribose + phosphate
deoxynucleoside + phosphate
Where are phosphodiester bonds formed?
Formed between the 3` OH position of one deoxynucleotide and 5` OH of the next
What bonds create the backbone of DNA?
repeating Phosphodiester bonds
Nucleic acids less than 50 nucleotides
Nucleic acids that are greater than 50 nucleotides in length
Chargoff's Rule
The abundance of deoxyadenosine always equaled that of thymidine and the abundance of deoxyguanosine always equalted that of deoxycytidine
How is the double helix stabilized?
The electrons of the adjacent base pairs interact, generating stacking forces and stabilize the helix in addition to the hydrogen bonds between the base pairs.
What are the different types of DNA conformations and which one is more prevalent?
A, B, and Z

The right handed B form is the most prevalent
What shape is the biological active form of DNA
The form is superhelical created by either underwinding or overwinding the double helix.
Small basic proteins containing large amounts of positively charged arginine and lysine that strongly binds to negative charged phosphates on DNA to package and order the DNA into structural units called nucleosomes
The complex that includes DNA, histones, and other proteins that makes up chromosomes
Consist of a segment of DNA wound around a histone protein core
Mitochondrial DNA
A small portion of eukaryotic DNA that is located in the mitochondria.

It encodes some, but not all of the mitochondrial proteins.
Structure of DNA
RNA chains are single-stranded
Lack the continuous helical structure of double-stranded DNA but does make stem-loop structure

Uses uracil rather than thymine

Sugar is a ribose
Small interfering RNA (siRNA)
Form of RNA that is effective in reducing the expression or completely silencing specific genes
What are the essential building blocks necessary for Purine synthesis?
Glycine, Glutamine, Aspartate and Tetrahydrofolate.
How does Purine synthesis differ from Pyrimidine synthesis?
Purines begin through the construct of the rings on a sugar while Pyrimidines do not.
What is the central intermediate for Purine synthesis?
Inosine Monophosphate (IMP)
What is the role of glutamine in Purine synthesis?
Glutamine donates its amine groups to make the central intermediate during Purine synthesis
What is the role of Glycine in Purine synthesis?
Glycine donates its carbon and nitrogen skeletons to make the central intermediate ring in Purine synthesis
What is the role of Aspartate in Purine synthesis?
Aspartate donates its carbon and nitrogen skeletons to make the central intermediate ring in Purine synthesis.
What is the role of tetrahydrofolate in Purine synthesis?
Tetrahydrofolate is a one carbon donor that participates in the formation of the central intermediate in Purine synthesis.
What is used to make the 5 membered ring of Purines?
Use glycine, glutamine and tetrahydrofolate and attach it to ribose
What is used to make the 6 membered ring of Purines?
Use aspartate's carbon skeleton and a carbon from tetrahydrofolate to finally make IMP.
Why is the addition of glutamine to PRPP a committed step?
PRPP has a pyrophosphate group at C1` which is an excellent leaving group, making the molecule high reactive, therefore, once the reaction occurs, the following molecules must be made.
How many ATPs are needed to make IMP?
4 ATPs
How do you make NDPs from NMPs?
Use kinases to add an additional phosphate at the 5` end
How do you make NTPs from NDPs?
Use nucleotide diphosphate kinases to add an additional phosphate at the 5` end
How does chemotherapy block purine synthesis?
Rapidly dividing cells, such as cancer cells, need purine synthesis to continue growing.

Use drugs which mimic the shape of a purine building block that can also inhibit an enzyme to block purine development
Protein enzyme that separates the DNA strands and unwind the parental duplex
Werner Syndrome
Rare disorder characterized by premature aging

Due to deficient helicases
Enzymes that wind/unwind DNA

Alter the linking number of DNA by three step process
1. Cleavage of one or both strands of DNA
2. Passage of segment of DNA through enzyme through this break
3. Resealing of DNA break
Topoisomerase Type I
Cleaves just one strand of DNA

Catalyzes the relaxation of a supercoiled DNA

Thermodynamically favorable
Topoisomerase Type II
Enzymes cleave both strands of DNA

Utilizes energy from ATP hydrolysis to add negative supercoils to DNA
What is the rate of DNA replication?
2000 bases are incorporated per second
Primer used in DNA replication
Short piece of RNA that is attached, usually to lagging strand of DNA, for DNA polymerase to catalyze replication
Primase catalyzes the synthesis of a short RNA segments, called a primer, complementary to a ssDNA template

Primase is activated by Helicase
In what direction does synthesis go in DNA replication?
Synthesis goes from 5` to 3`
DNA polymerase III

Primary enzyme complex involved in prokaryotic DNA replication

Catalytic prowess: 1000 nucleotides are added per second compared to 10nucleotides/sec for DNA polymerase I (It is 1000 times faster)

This is due to processivity

Has proofreading capabilities that correct replication mistakes by means of exonuclease activity (3` --> 5`)
Measure of the average number of nucleotides added by DNA polymerase enzyme per association/disassociation with the template
DNA polymerase I
an enzyme that participates in the process of DNA replication in prokaryotes and also removes RNA primer

Possesses three enzymatic activities:
1. 5' -> 3' (forward) DNA polymerase activity, requiring 3' primer site and template strand
2. 3' -> 5' (reverse) exonuclease activity that mediates proofreading
3. 5' -> 3' (forward) exonuclease activity mediating nick translation during DNA repair
Okazaki fragments
Synthesis of DNA that is produced in a 5` to 3` direction away from the fork in pieces, that is eventually joined together
DNA ligase
Enzyme that catalyzes the formation of phosphodiester bonds between the 3` hydroxyl group at the end of one DNA chain and the 5`-phosphate group at the end of the other strand.

Uses ATP in eukaryotes

Uses NAD+ in bacteria
What is the error rate of base-pairing?
Error rate is about 10^6

If proofreading activity is removed, the error rate increases to about 10^3
What are some of the differences between bacterial and eukaryotic DNA replication?
Replication is continuous in bacterial cells.
In eukaryotic cells it is done in phases.

Bacterial chromosomes have a single point of origin.
Eukaryotic chromosomes have multiple points of oritins at which replications begins.
Poymerase δ
The major eukaryotic enzyme involved in DNA replication on leading and lagging strands

Exonuclease Activity goes 3` to 5`
Single stranded binding proteins
Prevents the single strands of DNA from reassociating during replication
Enzyme that adds DNA sequence repeats ("TTAGGG") to the 3' end overhang of DNA strands in the telomere regions

Acts as a RNA-dependent DNA polymerase (like reverse transcriptase) because it contains both proteins and RNA

Once long, the primase enzyme can bind and synthesize the complementary strand
Xeroderma pigmentosum
Genetic disorder of DNA repair in which the ability to repair damage caused by ultraviolet (UV) light is deficient
Mismatch repair
Parental DNA strands contain methyl groups on adenine bases in specific sequences

During replication, the newly synthesized strands are not immediately methylated

Unmethylated strand is repaired
Not carcinogenic until it is oxidized

Covalently binds to guanine residues in DNA, interrupting hydrogen bonding in G-C base pairs

Produced distortions of the helix
Thymine dimer
UV light can cause two adjacent pyrimidines to form a covalent dimer
When the free ends of DNA at break points reseal with free ends of a different chromosome
Segments of DNA that can move from their original position in the genome to a new location

Found in all organisms
Reverse Transcriptase
An enzyme that uses a single-stranded RNA template to make a DNA copy

Found in retroviruses
Complementary DNA

The DNA copy of the RNA produced by reverse transcriptase
Synthesis of RNA from a DNA template

Catalyzed by a large enzyme called RNA polymerase
RNA polymerase
Catalyzes the initiation and elongation of RNA chains
Which strand of DNA is used for transcription?
Copying takes placed from complementary (template/antisense) strand.

Therefore the mRNA has the same sequence of genes as the coding (sense) strand
Why type of bonds are created by DNA polymerases and RNA polymerases
Forms ester bonds between nucleotides that base-pair with the complementary nucleotides on the DNA template
Difference between RNA and DNA polymerase
Unlike DNA polymerase, RNA polymerase can initiate the synthesis of new chains in the absence of primers

RNA polymerase also lacks 3` to 5` exonuclease proofreading activity
Bacterial RNA polymerase
Single RNA polymerase transcribes DNA to generate all forms of RNA (mRNA, rRNA, and tRNA)
Eukaryotic RNA polymerase
Three different RNA polymerases involved in transcription.
RNA Polymerase I
Eykaryotic RNA polymerase that produces most of the rRNA.

It is insensitive to the effects of alpha-amanitin
RNA Polymerase II
Eukaryotic RNA polymerase that produces mRNA

Strongly inhibited by alpha-amanitin, even under low concentrations of poison
RNA Polymerase III
Eukaryotic RNA polymerase that produces small RNAs like tRNA and 5S rRNA

Inhibited by alpha-amanitin only under high concentrations
Poison that binds tightly to RNA polymerase II.

Blocks the elongation phase of RNA synthesis

Higher concentrations of the poison also inhibits RNA polymerase III
Cis acting activating regions
The promotor region is on the same strand of DNA that is being transcribed
Trans acting proteins
Proteins that bind to promotor DNA sequences. They are not on the same strand.
Promotor region
Areas that are on the DNA that shows which genes to transcribe and where RNA polymerase should start transcribing them
Two basic markers for where transcription should start in Prokaryotes
TATA/Pribnow box (-10)

-35 base pair consensus sequence
How is frequency of transcription controlled
Controlled by regulatory sequences within the promotor and enhancers
Promotor-proximal elements
Interact with proteins that stabilize RNA polymerase binding to the promotor to help with rapid transcription
Eukaryotic promotors
TATA box present, but is more downstream (-20 to -30)

G-C rich sequence found at -40

CAAT box at -110
General Transcription Factors
AKA Basal factors

Bind to TATA box and facilitate the binding of RNA polymerase II to transcribe mRNA
Unwinds DNA for either transcription or DNA repair

Acts as an ATP-dependent DNA helicase
Transcription factors that act in the site of promotion
These generally work on many genes
Transcription factors that act at specific genes
Are gene specific transcription factors
What signals terminate transcription
Formation of hairpin loop on RNA preceding a number of Uracil residues

Rho factors associate with RNA polymerase and leads to termination
Rho factors
Rho factors are ATP-dependent helicases that associate with RNA polymerase and leads to termination

It pulls RNA away from the RNA polymerase
What leads to the formation of hairpin loops?
Palindrome sequences cause the RNA to fold back on itself forming the loop
A region of DNA that controls the process of transcription
Lac Operon
Repressor protein, encoded by promotor, blocks the promotion of genes to be transcribed. It prevents RNA polymerase from transcribing these genes

An inducer binds to the repressor protein, causing it to dissociate.

RNA polymerase can now recognize the promotor for genes.
A region of DNA that encodes a single polypeptide chain
Repressor Protein
Repressor protein, encoded by promotor, blocks the promotion of genes to be transcribed. It prevents RNA polymerase from transcribing these genes
A stimulator that binds to the repressor protein of lac operons, causing it to dissociate
Capping modification
Synthesized by RNA polymerase II

5` terminal end is modified such that a GTP is attached in a 5` to 5` linkage and methylation occurs at guanine and 2`C of ribose

It decreases the rate of degradation and serves as a recognition site for the binding of mature mRNA to ribosomes for translation
Poly(A) Tail
Modification of RNA where 250 adenine nucleotides are added to the 3` end by poly(A) polymerase
Removal of Introns
Modification of pre-mRNA where large regions are excised and the exons are spliced together.

It is carried out by spliceosomes
Assemblies of proteins and small RNA molecules (Ribozyme) that carry out splicing of Exons
Stop Codons
Degenerative genetic code
More than one codon can code for the same amino acid
Codons that specify the same amino acid
Point mutation
A single base change
Silent mutation
A change that specifies the same amino acid
Missense mutation
A change that specifies a different amino acid
Nonsense mutation
A change that produces a stop codon
Insertion mutation
An addition of one or more bases
Deletion mutation
A lost of one or more bases
Frame-shift mutations
Subclass of insertion/deletion mutation

When bases are added or removed, it causes a shift in the reading frame, therefore all the codons will be shifted
Loop on tRNA that is complementary to mRNA
5` end of tRNA
Phosphorylated area

Has an activated amino acid
3` end of tRNA
Activated amino acid is attached to the -OH group of 3`` end
Wobble Hypothesis
Some tRNA molecules can recognize more than one codon
Base found in anticodon (it is the first base of anticodon to match last base in codon)

Allows wobble base-pairs to form

It can bind to adenine, cytosine or uracil

Therefore, one tRNA molecule can base pair with three different codons
Aminoacyl-tRNA synthetases
Enzymes that attach amino acids to their tRNA

Twenty different enzymes exist, one for each amino acid
dHow does aminoacyl-tRNA synthetases chose tRNA partners?
Acceptor stem - position that contains amino acid at the end of the loop
Other Unique features like unusual bases and placement of bases
Ribosomal subunit
Large subunit (50S)
Small subunit (30S)
Together form 70S
Initiation of Translation
Involves the formation of a complex composed of methionyl-tRNAiMet, mRNA and a ribosome

Initiator tRNA binds to 30S part of ribosome. GTP and initiation factors are attached to activate this.

mRNA binds
P Site
Peptidyl site on ribosome
A site
Aminoacyl site on the ribosome
Shine-Delgarno sequence
Protein synthesis begins with the interaction of 30S subunit and mRNA through this sequence.
Differences between Eukaryotic and Prokaryotic Initiation of Protein Synthesis
1. Binding of mRNA to small ribosomal subunit is through 5` cap of mRNA
2. First amino acid is methionine
3. Initiation factors are elFs
4. Ribosomes are 80S (40S and 60S)

1. Binding of mRNA to small ribosomal subunits is through Shine-Dalgarno sequence
2. First amino acid is formyl-methionine
3. Initiation factors are IFs
4. Ribosomes are 70S (30S and 50S)
rRNA of large ribosomal subunit catalyzes the formation of the peptide bond in translation of mRNA
Uncharged tRNA moves from the P site and is release from the ribosome
Ribosomal release factor
A protein that resembles tRNA that recognizes stop codons and terminates translation
A group of Ribosomes that simultaneously translate a single mRNA
Bind to Nascent Polypeptide chains that protect against misfolding.
Post-translation modification
Initial methionine is removed by protease

Cleavages of proteins to activate (proinsulin to insulin)

Modified through phosphorylations, methylations, carboxylations, etc to change the behavior of the protein
Proteins synthesized on polysomes in the cytosol
After they are release from ribosomes, they remain in the cytosol, they they carry out their functions
Proteins synthesized on cytosolic ribosomes
Enter organelles, such as mitochondria or nuclei
Proteins synthesized on ribosomes bound to RER
Proteins are destined for secretion or for incorporation in organelles or cellular membranes
Chromatin Remodeling
Positive charge from amino group of lysine is acetylated, removing its charge, thereby reducing the electrostatic forces between histones and DNA.

Makes it easier for DNA to unwind
Methylation of 5` cytosine
Means of regulation of DNA to turn transcription and therefor translation off.
Antiobiotic that inhibits the peptidyl transferase activity of the 50S ribosomal subunit in prokaryotes