Genetic Code and Translation

Genetic Code

Learning Goals

• Explain how the redundancy of the genetic code can provide protection from mutation.

Information Flows from Genes to Proteins

• Proposed model by Franklin Crick and James Watson called “The central dogma of molecular biology.”

• Necessity of a "dictionary" to translate nucleotide language into amino acid language.

The Genetic Code

Codon Structure

• First position of codon (5' end)

• Second position of codon

  • U, C, A, G

  • Examples:

    • 5' end Codon: UCU, UAU, UGU

    • Amino Acids:

    • UUC = Phe, UCC = Ser, UAC = Tyr, UGC = Cys

• Third position of codon (3' end)

  • Examples:

    • UAA (Stop), UAG (Stop), UGA (Stop)

    • AUG = Met (Initiation codon)

Encoding Genetic Information

Properties of the Genetic Code

• The structure of DNA indicates that the sequence of amino acids in a polypeptide is dictated by the sequence of nucleotides in DNA of a gene.

• The information in a gene is represented in the form of a genetic code.

• Codons for amino acids are non-overlapping triplets of nucleotides.

• Distinction between overlapping and non-overlapping genetic codes.

Overlapping vs Non-Overlapping Genetic Codes

• Overlapping Code:

  • Example:

    • AUU, UUG, UGC

    • Multiple aa (amino acids) are encoded from a single nucleotide sequence.

• Non-Overlapping Code:

  • Example:

    • AUU, GCU, CAG

    • Each nucleotide belongs to only one codon.

Identifying the Codons

• Determined via transcription of artificial mRNAs.

• The genetic code is nearly universal; present in all organisms.

• The first two codon bases for a particular amino acid are invariant, while the third can vary.

• The universal decoder chart lists the 64 possible mRNA codons and their corresponding amino acids.

Characteristics of the Genetic Code

• Triplet Code: A triplet of nucleotides (codon) encodes one amino acid.

• Continuous Code: RNA is read continuously in groups of three nucleotides.

• Non-Overlapping: Each nucleotide is part of only one codon.

• Unambiguous: Each codon specifies one and only one amino acid.

• Nearly Universal: Almost all organisms share the same genetic code, allowing for mRNA from any organism to be translated correctly in other cell extracts (e.g., plants, animals, bacteria).

• Degenerate Code: 18 amino acids are encoded by multiple codons (synonymous codons); exceptions are methionine (AUG) and tryptophan (UGG).

• Start and Stop Codons:

  • Start codon: 5'-AUG-3' (Methionine).

  • Stop codons: 5'-UAG-3', 5'-UAA-3', and 5'-UGA-3' (terminate translation).

• Reading direction of mRNA: 5' to 3'.

• Polypeptide chain construction proceeds from N-terminus to C-terminus.

Genetic Evidence for a Triplet Code

• Normal Gene Example:

  • DNA: ACG TCA TAT CCG CAT ACC GAG

  • Amino Acids: Thr Ser Tyr Pro His Thr Glu

• Mutations:

  • Mutation +1 nucleotide:

    • DNA: ACG GTC ATA TCC GCA TAC CGA G

    • Amino Acids: Thr Val lle Ser Ala Tyr Arg

  • Results in inactive protein.

  • Mutation +2 nucleotides:

    • DNA: ACG GAT CAT ATC CGC ATA CCG AG.

    • Amino Acids: Thr Asp His lle Arg lle Pro

  • Results in inactive protein.

  • Mutation +3 nucleotides:

    • DNA: ACG GAC TCA TAT CCG CAT ACC GAG

    • Amino Acids: Thr Asp Ser Tyr Pro His Thr Glu

  • Results in active protein.

Role of Poly-U Template in Translation

• UUU encodes Phenylalanine:

  • Poly-U nucleotide template translates into a peptide sequence of phenylalanine (Phe).

  • Depiction of ribosomes, amino acids, and tRNAs involved in the process.

Translation

Learning Goals

• Describe the role of transfer RNAs (tRNAs) in protein synthesis.

• Outline the process of translation—from initiation through termination.

Overview of Translation

• Initiation: Ribosome binds mRNA at start codon.

• Elongation: Growing polypeptide chain develops.

• Termination: Encountering a stop codon (UAA, UAG, UGA) triggers release of the polypeptide and disassembly of the ribosome.

Basic Model for Protein-Encoding Gene Expression

Part 1

• Gene structure including:

  • 5' UTR

  • Open reading frame where translation occurs

  • 3' UTR

• Transcription is mediated by RNA polymerase.

Part 2

• Translation:

  • Involves ribosomes and tRNAs to create the polypeptide chain from the mRNA template.

  • Translation initiation requires binding of the small ribosomal subunit to the mRNA and assembling a functional complex.

Decoding the Codons: The Role of Transfer RNAs

Structure of tRNAs

• tRNAs are essential adaptors that decode information in mRNA.

• Structural characteristics:

  • Roughly the same length and shape, about 75-90 nucleotides long.

  • All mature tRNAs possess the triplet sequence CCA at the 3' end.

  • Two-dimensional representation shows cloverleaf form of yeast tRNA.

Wobble Hypothesis

• The third position of the codon permits variability; tRNAs can recognize more than one codon due to this phenomenon.

• Pairing flexibility:

  • U of the anticodon can pair with A or G of the mRNA

  • G of the anticodon can pair with U or C of the mRNA

  • I (inosine) of the anticodon can pair with U, C, or A of mRNA codon.

tRNA Charging

• Energy-required process linking amino acids to respective tRNAs through covalent attachment at CCA sequence (3' end).

• Carried out by aminoacyl-tRNA synthetases, which are specific to each amino acid.

• Two-step reaction process with proofreading capability (error rate ~ 1 per 10,000).

Translating Genetic Information: Initiation

• Protein synthesis is complex and similar across prokaryotes and eukaryotes.

• Translation phases: Initiation, Elongation, Termination.

Ribosomes

• Bacterial Ribosomes (70S):

  • Large subunit: 50S

  • Small subunit: 30S

• Eukaryotic Ribosomes (80S):

  • Large subunit: 60S

  • Small subunit: 40S

• Structure includes rRNA and proteins, varying between prokaryotes and eukaryotes.

mRNA Structure

• mRNA composition includes:

  • 5' untranslated region

  • Coding region protected by 5'-cap and poly(A) tail for stability against exonucleases.

• Shine-Dalgarno sequence:

  • Located upstream of AUG start codon for bacterial translation initiation.

  • Binding to rRNA in the small ribosomal subunit facilitates initiation.

Eukaryotic Translation Initiation

• Three steps incorporated:

  • Small subunit binds to the mRNA (at AUG).

  • Recruitment of aminoacyl-tRNA.

  • Assembly of initiation complex.

• At least 12 initiation factors (more than 25 polypeptide chains) are required for this process.

• 43S preinitiation complex must locate the 5' end of mRNA.

Elongation and Termination

Elongation Phases

• Aminoacyl-tRNA Selection:

  • GTPase protein EF-Tu assists in binding the second aminoacyl-tRNA to the A site.

• Peptide Bond Formation:

  • Ribosome catalyzes incorporation of approximately ten amino acids per second into the growing polypeptide chain.

• Translocation: Treadmill-on small subunit choreographed movement allowing advancement of ribosome along mRNA strand.

Termination

• Occurs at stop codons (UAA, UAG, UGA)— requires release factors that recognize these signals.

• The ribosome and mRNA dissociate; completed polypeptide is released.

Quality Control

• Nonsense mutations introduce premature termination codons in around 30% of inherited human disorders.

• Cells possess a mechanism for surveillance to detect premature termination in mRNA sequences.

Polyribosomes

• Complex of multiple ribosomes attached to a single mRNA strand for simultaneous translation, increasing protein synthesis efficiency.

Chaperone Proteins

Action During Translation

• Chaperones assist in protein folding and transport.

• Heat shock proteins (Hsp70) facilitate transitions through partially folded intermediates.

• Transfer to chaperonins provides protective conditions for proper protein folding completion.

Protein Isomerases

Peptidyl Prolyl Isomerase

• Enzymatic facilitation of switch between cis and trans conformations of proline residues within polypeptides, impacting protein folding dynamics.