BSCI331 Exam 2

what does the central dogma states

DNA → RNA → Protein

How does DNA go to RNA

RNA synthesis also called transcription

How does RNA go to protein

Protein synthesis also called translation

Does the central dogma refer to all biological systems?

No, there are exceptions such as Prions which are misfolded proteins which peer pressure normal proteins to fold weird without DNA transfer

what is the point of RNA transcription

To get the double stranded DNA into a singe stranded RNA

the strand that the RNA is complementary to is called?

The template strand on the DNA

What direction is RNA read in on the template strand

RNA reads the template strand 3 to 5 but synthesizes the new RNA molecule 5 to 3

How does RNA code look like?

ATCGTGCAT

UAGCACGUA

what enzyme reads the DNA in transcription

RNA polymerase

how does RNA polymerase (eukaryotes/prokaryotes) know where to start transcription?

recognizing and binding to promoters

how does RNA polymerase bind to promoters in eukaryotes since it is more difficult?

RNA polymerase is able to bind to general transcription factors (GTF) first and then be loaded onto the adjacent promoter sequence

How is DNA held together in chromating for transcription

As chromatin is looser it allows for transcription and gene expression

what enzyme can we find near the GTF that aids in chromatin opening

histone-modifying enzyme and chromatin remodeling complex

what is the first stage of RNA transcription?

Initiation — where RNA polymerase binds to a DNA’s gene promoter to start RNA synthesis

what is the second step of RNA synthesis/transcription called?

elongation — where the enzyme RNA polymerase moves along the DNA template strand synthesizing RNA

what proteins are required in the elongation of RNA?

elongation factors proteins using ATP hydrolysis

when does elongation end when running along the template strand?

in prokarytes this would be when the RNA polymerase hits the terminator sequence and in eukaryotes transcription ends when it hits a polyadenylation sequence

Most eukaryotic RNA needs what after?

post-transcriptional modifications before it becomes functional

what is the first mRNA post transcription modification after it leaves the RNA polymerase

5’ end capping — the addition of a 7-methylguanosine (m7G) cap

what does the 7 methylguanosine cap do?

it marks the RNA as a messenger RNA

marks the 5’ end as an “intact” end

binding site for the ribosome allowing it to bind to the mRNA

what characterizes the 5’ cap linkage

the 5’ cap is bound to the 7 methylguanosine via 3 phosphates and it is 5’ — 5’ bound

what is pre-mRNA processing?

it is the modification of newly transcribed precursor messenger RNA (pre-mRNA) into mature mRNA

what are introns and exons?

Exons are protein-coding nucleotide sequences in DNA that are transcribed

Introns are non-coding intervening sequences that interrupt coding regions

how are introns cut out so exons can be transcribed?

cut out in RNA splicing by using spliceosomes

what is the spliceosome made of?

snRNPs

snRNAs

some proteins

how does the spliceosome know where to splice?

there are sequences recognizable at the intron and exons boundaries

is the 5’ cap necessary for RNA splicing to occur

Yes it happens before splicing

when does RNA splicing occur?

during or immediately after the transcription of DNA → pre-mRNA

what is alternative splicing

it is a type of RNA splicing in which some genes are includes and excluded (exons) allowing different isoforms of the mRNA to be produced as different proteins

why is it important to carry around so many introns

gives the cell proteins diversity through alternative splicing without needing more DNA

example of a-tropomyosin gene alternative splicing

One DNA sequence but when it is transcribed we can pick and choose to cut out different introns from the gene to get many different forms of muscles

what is added to the pre-mRNA after RNA splicing

the poly-A tail is added to the 3’ end (~200 A’s are added)

What role does the poly-A tail play

allows the cell to recognize that this is a valid mRNA and it can be exported from the nucleus

where does all of the RNA synthesis and processing occur

in the nuclues

where does proteins synthesis occur

in the cytosol

what characteristics are required for pre-mRNA to finally become mRNA

5’ cap (polyadenylation tail)

Splicing is done

3’ cap (poly-A tail)

what is the breakdown of the RNA’s look like

mRNA = 5%
rRNA = 80%

what do mRNA’s even do?

messenger RNAs and used for translation, turning DNA into proteins

what do rRNA’s do?

they are ribosomal RNA’s and are used to form the structure of ribosomes and catalyze proteins structure

what do tRNA’s do?

transfer RNA used as a plug between mRNA and protein synthesis

what do snRNAs do?

they are used in RNA splicing along with snRNPs

once mRNA is transferred to the cytosol what can we do?

translation of mRNA to Proteins

Is translation a 1:1 communication

No, there are only 4 bases and 20 amino acids so there is some overlap

what is a codon

a set of 3 amino acids, there are more codons than amino acids, so amino acids have multiple corresponding codons

what is a degenerate genetic code?

An instance when multiple codon triplets can encode the same amino acid, providing redundant coding

how does the tRNA fit in the central dogma

tRNA recognizes the mRNA on one side and the other recognizes amino acid language

at what end does the amino acid chain begin in the tRNA

3’ end

how is the tRNA structured

contains a loop called with an anticodon that is complimentary to the amino acids codon

what does the anticodon do?

the 3 nucleotide sequence on the tRNA molecule that base-pairs with a complementary messenger RNA (mRNA) codon during protein synthesis

how are amino acids attached to the tRNA?

they are coupled via the aminoacyl-tRNA synthetase (each has it’s own — 20)

what are the steps of amino acids being attached to the tRNA

ATP is hydrolyzed by conjugation to AMP and activated

The amino acid is transferred from AMP to tRNA, resulting in a high-energy bond between the tRNA and the amino acid in the 3’ end

Then the synthetase proofreads for accuracy

photo of amino acid being attached to tRNA

amino acid becomes adenylated using ATP → AMP and then it is transferred to the high energy bond with tRNA making aminoacyl-tRNA on the 3’ end

which direction is protein synthesis added?

N→C terminus

amino acid is added to last tRNA that was added

diagram of new aminoacyl-tRNA replacing the last one when the protein residues increase

what does the rRNA do in translation?

they catalyze the peptide bonds and build the amino acids

what direction does the ribosome read mRNA?

5’ to 3’ reading 3 bases (codon) at a time

each mRNA has how many possible reading frames?

3 — so it is important for it to stay consistent and maintain the same reading frame we start with

what is actually reading the mRNA when it comes to the cytosol?

Ribosome

what brings the amino acids?

tRNA

What makes proteins?

Ribosome

where does the ribosome know when to start reading the mRNA?

the first AUG sequence (encodes for Met)

why does AUG (code for Met) use a special tRNA initiator?

to distinguish the start of protein synthesis from internal methionine additions during translation

what is the initiation process of translation consist of?

the assembly of the ribosome, mRNA, and initiator tRNA to begin protein synthesis and start with Met

what are the elongation factors called in prokaryotes and eukaryotes?

Ef-tu anf Ef-G

EF1 and EF2

how does EF-tu/EF1 work?

EF-tu/EF1 binds ATP to the aminoacyl-tRNA and brings it to the A-site and then it checks the codon-anticodon match

Proofreading step after:

If match is correct → GTP is hydrolyzed → EF-Tu/EF1 leaves

If match is wrong → tRNA falls off before GTP hydrolysis

EF-Tu uses GTP as a “checkpoint” to ensure correct tRNA binding

how does Ef-G/EF2 work?

Ef-G/EF2 work by using GTP to physically move the ribosome after the peptide bond begins to form

Translation elongation diagram

how do we end translation?

termination codon (UAA, UAG, UGA)

are there multiple ribosomes translating a singular mRNA at once?

Yes it is called a polyribosome which is a complex of multiple ribosomes

is the mRNA in a loop when it is being translated?

Yes

How does mRNA stay in a loop during translation?

5’ end is bound by eIF4E and 3’ end has poly-A binding protein (PABP)

between the elF4E and the 3’ polytail end it is bound by eIF4G which connects them

how do antibiotics work?

they work by blocking protein synthesis, if the rRNA of bacteria is blocked it will not be able to grow and the proteins will stop being synthesized

how do antibiotics not target us?

bacterial ribosomes are 70s and we are 80s

however since the mitochondria was phagocytized may still effect mitochondrial ribosomes

examples of antibiotics

Cycloheximide: prevents ribosome translocation by freezing it in place preventing the amino acid to form

Puromycin: larps as a tRNA so when it adds on after hydrolyzing GTP it causes the entire ribosome mRNA contraption to fall apart

when you add one nucleotide or one amino acid how much energy anre you using?

~2 ATP

what does tRNA and EF-Tu / EF-G use?

ATP and GTP

protein synthesis is very expensive

what does having protein tell us?

3D structure

Function

Where it goes in the cell

what does having mRNA tell us?

Protein sequence (codons)

Start/stop signals

Stability info

what does having DNA tell us?

Everything in mRNA

plus

Promoters (start transcription)

Terminators

Introns (regulation, splicing info)