tRNA, mRNA & Protein Synthesis

• Nucleic acid sequence alterations result in improper amino acid insertion into polypeptide chains, potentially causing disease or death.
• Translation converts mRNA information into new proteins, requiring energy from cleaved phosphoanhydride bonds.

Ribosomes and Protein Assembly

• Ribosomes are large ribonucleoprotein particles coordinating mRNA and tRNA interaction during protein synthesis.
• Ribosomes are products of ribosomal genes.
• The proteome encompasses all proteins produced by a cell at any given time.

Ribosome Structure

• Ribosomes consist of small and large subunits.

Prokaryotic Ribosomes

• Bacterial cells have approximately 20,000 ribosomes, 25% of the cell mass
• Escherichia coli ribosome sedimentation coefficient: 70 S
• Contain three rRNA molecules and up to 83 proteins
• Prokaryotic Ribosomes consist of about 65% RNA and 35% protein
• The sedimentation coefficient measures molecule rate in an ultracentrifuge in a less dense solvent.
• Sedimentation coefficients are measured in Svedberg units (S).

Prokaryotic Ribosome Subunits

• 30 S subunit
• Site for genetic information decoding and contains proofreading mechanisms
• Contains 16 S rRNA and 21 proteins
• 50 S subunit
• Smaller rRNAs: 5 S (120 ribonucleotides) and larger rRNAs: 23 S (~2900 ribonucleotides), and 3-35 proteins
• The 50 S subunit provides peptidyltransferase activity

Eukaryotic Ribosomes

• Eukaryotic ribosome size: 80 S (4.2 MDa)
• 40 S Subunit
• Contains 18 S rRNA (1900 bases) and 33 proteins
• 60 S Subunit
• Contains 5 S rRNA (120 bases), 5.8 S rRNA (160 bases), 28 S rRNA (4800 bases), and 50 proteins

Mitochondria and Chloroplasts

• Mitochondria and chloroplasts contain ribosomes similar to prokaryotes.
• Structure is sensitive to translation antibiotic inhibitors.

Nucleolus and Ribosomes

• The nucleolus is functionally specific in the cell nucleus, where ribosomes are synthesized.
• Human rRNA genes number 200 per haploid genome and are transcribed by RNA polymerase I, forming 45 S rRNA precursors.
• rRNA precursors package with ribosomal proteins from the cytoplasm, cleaving three of the rRNA subunits.
• Three of the four rRNA subunits transfer to the nucleus and release the 5S RNA to form functional ribosomes

Small Nuclear RNAs

• Small nuclear RNAs are RNA molecules that bind specifically with nuclear ribonucleoprotein particles (snRNPs).
• Small nuclear RNAs play important roles in RNA molecule post-transcriptional modification.
• Small nuclear RNAs base-pair with pre-mRNA and during RNA splicing.
• Small nuclear RNAs are made of non-coding RNA and participate in splicing.

Small Nucleolar RNAs

• Small nucleolar RNAs aid in processing pre-rRNAs and ribosome assembly.

Medical relevance

• Chemical compounds inhibit transcription or translation in cancer therapies and as poisons
• Cancer effectors involving growth factors, proteins, influence transcription and translation.

Genetic Code

• System of RNA sequences specifies amino acids during translation

Codons

• Genetic words composed of 3 nucleotide bases (adenine, guanine, cytosine, uracil)

Codon Properties

• Codons are arranged 5' to 3'.
• The four nucleotide bases produce 64 different combinations.

Translating Codons

• Tables translate codons and determine which amino acids an mRNA sequence codes for.
• For example, the codon 5'-AUG-3' codes for methionine.
• AUG is the initiation (start) codon for translation.
• Sixty-one of the 64 codons code for the 20 common amino acids.

Termination Codons

• Three codons (UAA, UAG, UGA) do not code for amino acids, but, rather, terminate codon.
• When termination codons appear, polypeptide synthesis stops.

Altering Nucleotide Sequences

• Changing a single nucleotide base (point mutation) yields different results include "silent" mutation.
• An example of silent mutation: If the serine (Ser) codon UCA changes to UCU, it still codes for Ser.
• Missense mutation: the new codon coding for a different amino acid.
• An example of a missense mutation: A different first base, changes the Ser codon UCA to CCA, coding for proline.
• The substitution of an incorrect amino acid is a "missense" mutation.
• Nonsense mutation: the new codon becoming a termination codon causing translation termination.
• Other mutations: These can alter expression or structure.
• Examples of other mutations: Trinucleotide expansion, Splice site mutations, and Frame-shift mutations

Genetic Code Characteristics

• The genetic code tends to be universal
• Differences can be found in the manner that it is translated

Redundant/Degenerate Code

• More than one codon specifies a single amino acid.
• Synonyms are codons designating the same amino acid.
• Synonymous codons usually differ only in the 3rd base of the codon

Unambiguous and Non-Overlapping Codes

• Each codon specifies no more than one amino acid

Nonoverlapping and Commaless

• If one or two nucleotides are shifted a frameshift mutations ensues in the amino acid that is coded
• If three nucleotides are shifted it has little to no effect on whether the protein has been translocated

Colinearity of Gene Product

• The product is a peptide specified by the sequence of expressed regions

Overlapping Genes

• Some viruses code for more proteins from their expressed portions

Translation components

• Amino acids, mRNA, tRNAs, ribosomes, energy sources, and enzymes are components required for translation

Amino Acids

• They have to be present for efficient codon-specifying action
• Absence causes translation to stop at that specified site

tRNA

• Need a specific type per amino acid
• They are also called adaptor molecules who can take the amino acid to the codon for that amino acid

Amino-Acid Attachment

• Attachment happens at the 3’ end of tRNA
• Carboxyl group from amino acids links to the hydroxyl of ribose as adenosine

Amino Acid Attachment Sites

• tRNA has a covalently attached amino acid, it's considered "charged"
• tRNA that isn't bound is "uncharged"

Aminoacyl-tRNA Synthetases

• Catalyzes attachment of amino acids to tRNAs
• Recognizes the amino acid and all tRNAs for that acid with specificity
• Catalyzes a two-step reaction, bonding an amino acid's carboxyl group to its corresponding tRNA's 3'-end

Energy Requirements

• Overall reaction needs adenosine triphosphate, cleaved to AMP and inorganic pyrophosphate
• synthetases are extremely specific, translating the genetic message with great accuracy which can remove incorrect amino acids

Anticodon

• Molecule with nucleotide sequence recognizing a particular codon
• Molecule is specified to grow and grow given each transfer

Ribosomal Subunits

• Play an important role in the structure with other steps as well as other components during the entirety of the translation process

tRNA molecules

• Binding sites A (Acceptor), P(Peptidyl), E (Exit) on ribosome that cover two adjacent codons molecules

Elongation

• Eukaryotes need additional ATP for this

Summary of Amino-Acid Activation

• Aminoacylated tRNAs are the link between the protein and its message
• tRNAs are covalently attached to a given amino acid

Adaptor Molecules

• aminoacyl-tRNA synthetases carry out the genetic code and implement all of it for you

Aminoacylation Steps

• Attachment of amino acids to the tRNAs
• Attachment also involves the amino acids binding to ATP

tRNA Properties

• The nucleotide sequence of the anticodon ensures that they have to be written from the 5’-end to 3’-end

Wobble Hypothesis

• The process by which trnas an recognize >a codon given the specification for any amino acid

Start Signals

• 30S requirements.
• mRNA strand what has to to be translated
• Factors which have to put the complex together

Antibiotics and Protein Synthesis

• Streptomycin prevents tRNA binding to the P site by binding to the 30S subunit
• Tetracycline prevents binding of aminoacyl-tRNA to the 30S subunit's A site.
• Chloramphenicol prevents prokaryotic ribosome from performing peptidyltransferase function.