Protein synthesis

transcription

• 1 of 2 DNA strands is used as a template for synthesising an RNA molecule - done by RNA polymerase

• occurs in nucleus

• steps:

  1. RNA polymerase unwinds DNA, exposing the bases

  2. RNA polymerase pairs up free RNA nucleotides to DNA nucleotides on template strands using complementary base pairing

  3. RNA polymerase links RNA nucleotides together w/ covalent bonds between pentose sugars and phosphate, forming continuous RNA strand

  4. RNA strand separates from DNA, DNA rewinds

hydrogen bonding in transcription

• bases in RNA/DNA pair up by hydrogen bonding

• easily broken to separate RNA transcript from template strand

conservation of DNA templates

• DNA very stable due to H bonding between bases and phosphodiester bonds between adjacent nucleotides

• stability of DNA means that genetic code isn’t prone to spontaneously breaking/changing

• DNA strands act as reliable templates for transcription over many generations

transcription and gene expression

• at specific times, some genes are expressed and some are not

• first stage in gene expression is transcription - producing an RNA copy of the base sequence of sense strand by transcribing template strand

• stage where genes can be switched on/off to match requirements of cell

translation

• taking the genetic code from mRNA and synthesising a polypeptide

• each base in polypeptide is coded for by one codon on mRNA (sequence of 3 bases)

• mRNA template comes from transcription, so translation occurs after

• occurs in cytoplasm

role of mRNA in translation

• occurs on ribosomes, which have 1 large subunit and 1 small

• steps:

1. mRNA binds to small subunit of ribosome. mRNA contains series of codons consisting of 3 bases, each codes for 1 amino acid

2. tRNA (transfer) present around ribosomes. each has an anticodon (3 bases) and carries corresponding amino acid to anticodon

3. 3 binding sites for tRNA on large subunit, only 2 ever bind at once. tRNA can only bind if has anticodon complementary to codon on mRNA. bases on codon and anticodon link by H bonds using complementary base pairing

4. amino acids carried by tRNA bond together through peptide linkage, forming dipeptide. 1 tRNA detaches with the dipeptide, and ribosome moves along mRNA to the next codon

genetic code

• codons are triplets to make sure they account for all the 20 diff types of amino acid

• degeneracy: there are more codons than needed for the 20 amino acids, so there are 2+ codons for most amino acids

• universality: 64 codons of genetic code have the same meanings in cells of all organisms

elongation of polypeptide chain

• ribosome moves along mRNA molecule 1 codon at a time

• peptide bond is formed between 2 amino acids (one on each tRNA)

• process continues until stop codon on mRNA is reached, so translation stop and polypeptide chain is complete

• amino acid chain then released from ribosome

mutations altering protein structure

• a point mutation = single base change

• single base changes one codon in mRNA, changing the amino acid synthesised from it

• one chained amino acid in polypeptide can cause radical changes in protein structure

• e.g sickle cell mutation

  • HBB gene codes for beta polypeptide of hemoglobin

  • single base change changed codon for 6th amino acid, causing sickle cell

direction fo transcription and translation

• transcription:

  • RNA polymerase adds the 5’ end of free RNA nucleotide to the 3’ end of growing mRNA molecule

  • occurs 5’ to 3’

• translation:

  • ribosome binds to mRNA near the 5’ end and moves along it towards the 3’ end

  • occurs 5’ to 3’

transcription factors

• adjacent to start of every gene is a section of DNA called a promoter that initiates transcription but isn’t transcribed itself

• RNA polymerase binds to promoter and starts transcribing in prokaryotes

• in eukaryotes, proteins called transcription factors first bind to promoter, then allowing RNA polymerase to bind

• a cell can switch on some genes and cause them to be transcribed while other genes are not

non-coding sequences

• coding sequences are transcribed and translated when a cell requires the protein they code for

• functions of non-coding sequences:

  • regulating gene expression

    • some base sequences are sites where proteins can bind that either promote/repress the transcription of an adjacent gene

  • introns

    • coding sequences are often interrupted by 1+ non-coding sequences

    • these introns removed from mRNA before it is translated

  • telomeres

    • repetitive base sequences at the ends of chromosomes

    • telomeres prevent important genes at the end of chromosomes from being lost everytime DNA is replicated

  • genes for tRNA and rRNA

    • transcription of these genes produces transfer RNA for translation and ribosomal RNA

post transcriptional modification

• eukaryotes modify mRNA after transcription before translation

1. removal of nucleotides from within transcript:

   • introns are removed from mRNA before it is translated

   • remaining parts of mRNA are exons, which are spliced together to form mature mRNA

2. addition of nucleotides to ends of transcript

   • RNA transcript modified by adding extra nucleotides to ends of mRNA to stabilise them and protect from digestion

   • 5 prime cap: modified nucleotide added to 5’ end of RNA

   • poly A tail: 100-200 adenine nucleotides added to 3’ end of RNA

alternative splicing

• some genes have many exons and different combos of them can be spliced together to produce different proteins

• increases total number of proteins that can be produced from organisms genes

start of translation

• sequence of events:

  1. initiator tRNA with anticodon AUG binds to small subunit of ribosome, carrying an amino acid

  2. small subunit of ribosome and initiator tRNA attach to 5’ terminal of mRNA and move along it until they reach start codon UAC

  3. anticodon of initiator tRNA and start codon form H bonds between their bases

  4. initiator tRNA binds to large subunit of ribosome binds at P site. E and A sites vacant

  5. tRNA with anticodon complementary to codon adjacent to start codon binds to A site. this tRNA carries amino acid that corresponds to the codon

  6. peptide bond forms between 2 amino acids from tRNAs in P and A sites

  7. cycle repeats elongating the polypeptide

tRNA binding sites

• A site: binding of a tRNA when it arrives carrying an amino acid

• P site: initiator tRNA binds here, normal tRNA moves here when it is carrying the growing polypeptide, polypeptide is transferred to the A site by formation of peptide bond

• E site: tRNA exits from here when it is no longer holding polypeptide

modification of polypeptides

• many polypeptides must be modified before they can function

• e.g the insulin gene is transcribe and then mRNA produced is translated into a polypeptide preproinsulin

• this is converted to proinsulin by a protease from rough ER which removes many amino acids

• proinsulin folds and disulfide bonds form between diff sections of polypeptide

• proteases in golgi remove more amino acids resulting in 2 chains attached by disulfide bonds

amino acids recycled by proteasomes

• damaged or unneeded proteins can be recycled into usable proteins

• proteasomes are organelles that digest selected proteins, releasing amino acids to be reused

• this process is essential for sustaining a functional proteome