Mod 6 Protein Synthesis
What are proteins?
-chains of amino acids
-proteins are synthesized into two main stages
-proteins determine the phenotype of an organism or a cell
How are proteins synthesized?
- DNA is transcribed into RNA through transcription
- RNA is then translated at the ribosomes to synthesis proteins
How are proteins synthesised in prokaryotes?
- prokaryotes such as bacteria, because there is no nucleus, RNA is immediately translated into proteins
How are proteins synthesised in eukaryotes?
- transcription takes place inside the nucleus
- the newly produced RNA is processed before leaving the nucleus and then is translated at the ribosomes in the cytoplasm
What did Archibald Garrod suggest?
-genes dictate phenotypes through enzymes that catalyze specific reactions
-he thought that the symptoms of an inherited disease reflect an inability to synthesize a particular enzyme
What is a Central Dogma?
-the central dogma of genetics is the flow of information in a cell
-it is always in one direction, DNA codes for RNA & RNA codes for protein
Central Dogma: phenotype & alleles
-an organism's genotype is determined by the sequence of bases in its DNA, while its phenotype is a product of the proteins it produces
-alleles of the same gene differ in their DNA sequence, the proteins produced by different alleles of the same gene frequently differ in their amino acid sequence
What are the exceptions to the central dogma:
-many genes code for the other RNA molecules that do not function as mRNA & are not translated into proteins such as tRNAs & rRNAs
-sometimes information flows in the opposite direction, from RNA back to DNA: this can be seen in some viruses such as HIV
RNA structure: How do DNA & RNA differ in their structure?
-RNA is a single-stranded molecule
-RNA contains the pyrimidine Uracil (U) instead of Thymine (T)
-RNA contains a ribose sugar whereas DNA contains a deoxyribose sugar
What is Transcription?
-the synthesis of RNA from a DNA template
-there are three types of RNA that are transcribed and involved in the process of translation
-these are messenger RNA (mRNA), transfer RNA (tRNA) & ribosomal RNA (rRNA)
What is Messenger RNA?
-transcription of most genes produces a messenger RNA which is translated at ribosomes to synthesize proteins
-messenger RNA takes the message from chromosome to ribosome using the DNA template
-the complementary mRNA molecule is synthesized according to base-pairing rules except that Uracil pairs with Adenine
-3 nucleotides make up one codon specifying one amino acid
termed as the genetic code
-mRNA has a 5' to 3' polarity/direction
What is Ribosomal RNA?
-ribosomal RNA is a part of ribosomes
-complementary base pairing binds mRNA & tRNA on the ribosomes
-rRNA consists of two pairs that function together: a large & small subunit
- the ribosomes are named based on their sedimentation rate
-eukaryotes: 60S & 40S (the subunits are made in nucleus)
-prokaryotes: 50S & 30S
What are Ribosomes?
-each ribosome has a binding site for mRNA & three binding sites for tRNA
-the A site (acceptor site) holds the tRNA with the next amino acid to be added to the chain
-the P site (peptide site) holds the tRNA carrying the growing polypeptide chain
-discharged tRNAs leave the site at E (Exit site)
What is Transfer RNA?
-the mRNA message is translated into proteins with the help of transfer RNA
-tRNA is used for picking up its designated amino acid, depositing it at the ribosome & returning to the cytoplasm to pick up another one
-molecules of tRNA are not identical: each carries a specific amino acid on the end
-each has an anticodon on the other end; the anticodon base pairs with a complementary codon on mRNA
-tRNA binds to the ribosome
Why can't proteins be translated directly from DNA?
1. to protect DNA
2. more copies of a protein can be made simultaneously since many RNA transcripts can be made from one gene
3. each RNA transcript can be translated repeatedly
What is RNA polymerase?
-separates the DNA strands & adds complementary nucleotides to the template strand
-one type of RNA polymerase is responsible for transcription prokaryotes but eukaryotes have three (I, II, III)
-RNA polymerase II used to synthesize mRNA
-assemble nucleotides in a 5' to 3' direction, a primer is not needed
What is the promoter?
- is a special sequence of nucleotides located at the start of a gene
-once found, RNA polymerase binds to the promoter region, unwinds DNA & begins transcription by attaching complementary nucleotides
The promoter: prokaryotes vs eukaryotes
-in prokaryotes, RNA polymerase is made up of the core enzyme that has the ability to synthesize RNA & a sigma unit
-sigma acts as a regulatory factor, guiding RNA polymerase to specific promoter sequences on the DNA template strand
-eukaryotes have a much more diverse & complex series of promoters
-most have a sequence called the TATA box, 30 base pairs upstream of that site
-transcription factors bind to DNA promoter, initiating transcription
What is a polyribosome?
-refers to the process when more than one ribosome is translating one mRNA molecule producing more proteins as required
-can be seen in prokaryotes due to the absence of a nucleus, they can transcribe & translate the same gene simultaneously
What are exons & introns?
-the protein-coding regions of eukaryotic genes, called exons are interrupted by noncoding regions called introns
-the final mRNA product will only have exons & no introns
-eukaryotic genes are therefore much larger than their corresponding mature mRNA
What is the splicing of RNA?
-the transcription of eukaryotic genes by RNA polymerase generates a primary RNA transcript that contains exons & introns
-introns are removed by splicing
-small nuclear ribonucleoprotein (snRNPs) for a complex called a spliceosome that removes introns & joins exons
Adding Caps & Tails to RNA Transcripts:
-primary RNA transcripts are further processed by adding a 5' guanine cap & a 3' poly Adenine tail
- the 5' cap serves as a recognition signal for the translation machinery
-the poly-A tail extends to the life of an mRNA by protecting it from degradation
What is the genetic code?
-the genetic instructions for a polypeptide chain are based on a triplet code, which is a series of nonoverlapping three-nucleotide sequences called a codon.
-all of the 64 codons make up what is termed the Genetic Code
-the position of a triplet on the DNA strand determines the corresponding position of an amino acid in the polypeptide chain
What is the redundancy of the genetic code?
-the genetic code is termed redundant, meaning the different codons can code for the same amino acid
-however, one codon will always code for only one amino acid
-tRNA can recognize more than one codon for a particular amino acid because the third base is relaxed, this is termed wobble
What are the components of translation?
-translation translates the language of mRNA into polypeptides
mRNA: directs amino acid sequence
tRNA: carries correct amino acids
rRNA + proteins: Ribosomes which have large & small subunits, E,P & A sites
A site: accepts the tRNA carrying an amino acid
P site: where peptide bonds form between amino acids
E site: where discharged tRNAs leave at the exit site
What are the two enzymes important in translation?
- Aminoacyl-tRNA Synhetase (AATS): attaches the correct amino acid to its tRNA
- Peptidyl Transferase: catalyzes the formation of peptide bonds between amino acids in the growing polypeptide chain using ATP & GTP for energy
What are the three steps Translation can be divided into?
- initiation
-elongation
-termination
What is the initiation of translation?
-at initiation, the small subunit of a ribosome recognises the 5' cap & attaches to the binding site of an mRNA
-once it recognises the start codon AUG, a charged tRNA carrying methionine attaches to the P site on the small subunit
-the binding of the large ribosomal unit with mRNA/tRNA & the small ribosomal subunit forms the initiation complex
What is the elongation of translation?
-charged tRNA carrying its specific amino acid enters the empty A site & binds to the mRNA codon
-the peptide bond is formed between amino acids that are located on the tRNAs in the P site & A site
-the elongation factor moves the mRNA down the ribosome one codon at a time; the tRNA located on the A site moves into the P site, & moves the P to the E site which- is termed TRANSLOCATION
-the A site is available to accept new aminoacyl-tRNA for binding
What is the termination of translation?
1. release factor binds to stop codon
2. polypeptide is released
3. Ribosome subunits separate
What are the post-translational modifications?
-signal regions in the peptide trigger its escort to specific regions by SRPs (signal recognition particles)
-proteins undergo post-translational modification
-folding determines its shape & function
-enzymes may add or remove a phosphate group: these changes can activate or deactivate a protein
What is the definition of a gene?
-the entire nucleic acid sequence needed for the synthesis of RNA sequence or functional polypeptide
What is the definition of a mutation?
-permanent alterations in the DNA sequence so that the sequence differs from what is found in most people
-can occur when an incorrect base is inserted during DNA replication
What are the two types of mutation?
1. Substitution- of one base for another ex. A for C, G for T
2. Insertion or deletion- of nucleotide base/s within the DNA sequence
What is the Missense Mutation? & what does it cause?
-caused by a change in a single nucleotide from T to A in the DNA template
-this changes the codon for amino acid 6 from glutamic acid to valine which causes aggregation of the mutant molecules instead of correct folding- resulting in distorted red cells
-can cause sickle cell disease
What is the Indel mutation?
-known as a frame-shift mutation
-nucleotide insertion or deletion
-changes the reading frame of the DNA sequence, so that codons are read incorrectly from the mutation onwards
-can cause nonsense mutation or missense mutation
What is a silent mutation?
-have no effect on the amino acid produced by a codon because of redundancy in the genetic code
-do not have apparent clinical consequences
What is a nonsense mutation?
- changes an amino acid codon into a stop codon, nearly always leading to a nonfunctional protein