(BIOCHEM) Chapter 7 - RNA and the Genetic Code

Central Dogma of Molecular Biology

DNA-transcription-RNA-translation-protein

What direction is mRNA synthesized?

5'--\> 3'

What direction does ribosome translate the mRNA?

5'--\>3' (N-terminus --\> C-terminus)

What are the three types of RNA?

mRNA, tRNA, rRNA

Messenger RNA

carries the information specifying the amino acid sequence of the protein to the ribosome

monocistrionic

\-each mRNA molecule translates into only one protein product

\-such is the case in eukaryotes

polycistrionic

\-an mRNA molecule may code for multiple proteins, depending on where translation starts

\-such is the case in prokaryotes

Transfer RNA

responsible for converting the language of the nucleic acids to the language of the amino acids and peptides

How are tRNA molecules activated with an amino acid?

aminoacyl-tRNA synthetase that requires two high-energy bonds from ATP; transfers amino acid to 3' end of the tRNA

Ribosomal RNA

synthesized in the nucleolus and functions as an integral part of the ribosomal machinery used during the protein assembly in the cytoplasm

Ribozyme

enzymes made up of RNA molecules instead of peptides

Codon

three letter word in mRNA, translated into an amino acid

Which direction are codons written in?

5'--\>3'

Start codon

AUG

Stop Codons

UGA, UAA, UAG

Degeneracy of genetic code

More than one codon codes for an amino acid

Wobble position

third position of the codon

two bases are usually the same - the third base in the codon is usually variable

two bases are usually the same - the third base in the codon is usually variable

used to protect against mutations in the coding regions of DNA

mutations in this position tend to be *silent or degenerate*

* there is no effect on the expression of the amino acid and therefore no adverse effects on the polypeptide sequence

Do silent or degenerate mutations affect the expression of the amino acid?

No

Point mutation

mutation occurs and only affects one of the nucleotides in the codon

Expressed mutations

point mutations that can effect the first or second nucleotide in the codon - changing the primary amino acid sequence of the protein

can be a missense or nonsense mutation

Missense mutation

mutation where one amino acid substitutes for another

Nonsense mutation (truncation mutation)

a mutation where the codon now encodes for a premature stop codon

Frameshift mutation

occurs when some number of nucleotides are added or deleted from the mRNA sequence

transcription

creation of mRNA from a DNA template

Newly synthesized mRNA strand is both \______ and \______ to the DNA template strand

antiparallel and complementary

How does RNA polymerase know where to start?

binds to specific areas of DNA called promoters

main player in transcribing mRNA

RNA polymerase II

Binding site of RNA polymerase II

TATA box; transcription factors help it locate and bind to promoter region of DNA

Three types of RNA polymerases

RNA polymerase I, RNA polymerase II, RNA polymerase III

RNA polymerase I

located in the nucleolus and synthesizes rRNA

RNA polymerase II

Located in the nucleus and synthesizes hnRNA and some snRNA.

RNA polymerase III

located in the nucleus and synthesizes tRNA and some rRNA

Numbering of bases

Positives are to the right, toward 3' end (upstream)

Negatives to the left, toward 5' end (downstream)

Where is the TATA box usually located?

-25 upstream

heterogeneous nuclear RNA

the initially synthesized mRNA strand containing both introns and exons

hnRNA must undergo three processes before leaving the nucleus

Splicing, 5' Cap, 3' Poly-A Tail

Splicing

the process of removing introns and reconnecting exons in a pre-mRNA; ligate coding sequences together (exons)

What does the splicing?

spliceosome - comprised of small nuclear RNA (snRNA) and small nuclear ribonucleoproteins (SnRNPs)

Where is the 7-methylguanylate triphosphate cap placed?

5' end

When is the 5' cap added?

During process of transcription

Function of 5' cap

recognized by ribosome as binding site and protects mRNA from degradation in the cytoplasm

3' Poly-A tail

protects mRNA from degradation; composed of adenine bases

Analogy for 3' Poly-A tail

like a fuse of a time bomb, the longer it is, the longer the mRNA will be able to survive before being digested in the cytoplasm

Alternative splicing

Splicing of introns in a pre-mRNA that occurs in different ways, leading to different mRNAs that code for different proteins or protein isoforms. Increases the diversity of proteins.

Where does mRNA exit the nucleus?

nuclear pores

What composes the ribosome?

Proteins and rRNA

Main function of ribosome

to bring the mRNA message together with the charged aminoacyl-tRNA complex to generate the protein

What are the three binding sites in the ribosome for tRNA?

A site (aminoacyl), P site (peptidyl), E site (exit)

Where does translation occur?

cytoplasm

Where does transcription occur?

nucleus

Three stages of translation

initiation, elongation, termination

Initiation (Prokaryotes)

small subunit binds to the Shine-Dalgarno sequence in the 5' UTR of mRNA

Initiation (Eukaryotes)

small subunit binds to the 5' cap structure; charged initiator tRNA binds to start codon within the P site of the ribosome

Initiation factors

assist in the binding of the large subunit and the small subunit to form the completed initiation complex

Order of sites in the ribosome during translation

APE

How many steps in elongation?

three-step cycle

A site

holds the incoming aminoacyl-tRNA complex

P site

holds the tRNA carrying the growing polypeptide chain

E site

where inactivated tRNA pauses transiently before exiting the ribosome

Elongation factors (EF)

assist by locating and recruiting aminoacyl-tRNA along with GTP, while helping to remove GDP once the energy has been used

release factor (RF)

binds to termination codon, causing water molecule to be added to polypeptide chain

What does the water molecule do in termination?

hydrolyze the completed polypeptide chain from the final tRNA

chaperones

Proteins that assist in protein folding during posttranslational processing

Modified by cleavage events (i.e. insulin)

cleaved from a larger, inactive peptide to achieve its active form

Phosphorylation

addition of phosphate group by protein kinases to activate or deactivate proteins; in eukaryotes most commonly seen with serine, threonine, and tyrosine

Carboxylation

addition of carboxylic groups, usually serve as calcium-binding sites

Glycolysation

addition of oligosaccharides as proteins pas through the ER and Golgi apparatus to determine cellular destination

Prenylation

addition of lipid groups to certain membrane-bound enzymes

Operon

a cluster of genes transcribed as a single mRNA

Jacob-Monod Model

operons contain structural genes, an operator site, a promoter site, and a regulatory gene

Structural gene

codes for protein of interest

operator site

A nontranscribable region of DNA that is capable of binding a repressor protein

promoter site

provides a place for RNA polymerase to bind

regulator gene

codes for repressor protein

What are the two types of operons?

inducible and repressible

inducible systems

the repressor is bonded tightly to the operator system and thereby acts as a roadblock

negative control

binding of a protein reduces transcriptional activity

How can the block be removed in an inducible system?

Inducer must bind to the repressor protein so that RNA polymerase can move down the gene

catabolite activator protein (CAP)

binds with cAMP and then binds to promoter region of operon to increase transcription

positive control

binding of a molecule increases transcription of a gene

Repressible systems

- allow constant production of a protein product; the repressor maintains inactivity until bound with a corepressor

What must bind to repressor to activate it?

corepressor

repressor-corepressor complex

when repressor binds the corepressor, can bind operator,

prevents transcription

transcription factors

transcription-activating proteins that search the DNA looking for specific DNA-binding motifs

Two recognizable domains of transcription factors

DNA-binding domain and activation domain

DNA-binding domain

binds to a specific nucleotide sequence in the promoter region or to a DNA response element to help in the recruitment of transcriptional machinery

activation domain

Allows for binding of several transcription factors and other important regulatory proteins

Ways for gene amplification

enhances and gene duplication

Enhancers

allows for control of one gene's expression by multiple signals

Gene duplication

can be duplicated in series; can be duplicated in parallel by opening gene with helicases and permitting DNA replication only of that gene

histone acetylases

involved in chromatin remodeling; acetylate lysine residues found in amino terminal tail regions of histone proteins

Acetylation

decreases positive charge on lysine residues and weakens interaction of histone with DNA, resulting in open chromatin conformation

Histone deactylases

remove acetyl groups, leading to chromatin condensation

DNA Methylation

involved in chromatin remodeling and regulation of gene expression levels in the cell

DNA methylases

add methyl groups to cytosine and adenine nucleotides; methylation associated with silencing of gene expression