Translation: The Molecular Biology of Protein Synthesis

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Translation: The Molecular Biology of Protein Synthesis

Overview of Gene Expression

Gene expression is the process by which genetic information encoded in DNA is used to produce functional proteins. This occurs through two main steps: transcription (DNA to RNA) and translation (RNA to protein). The central dogma of molecular biology describes this flow of information: DNA → RNA → Protein.

Transcription: Synthesis of RNA from a DNA template.
Translation: Synthesis of a polypeptide (protein) from an mRNA template.

Central dogma: DNA to RNA to protein Gene structure and transcription/translation overview

The Structure and Properties of Amino Acids

Proteins are composed of 20 common amino acids, each with unique chemical properties. Amino acids are classified based on their side chains as nonpolar, polar, acidic, or basic.

Nonpolar: Glycine, Alanine, Valine, Leucine, Isoleucine, Methionine, Tryptophan, Phenylalanine, Proline
Polar: Serine, Threonine, Cysteine, Tyrosine, Asparagine, Glutamine
Acidic: Aspartic acid, Glutamic acid
Basic: Lysine, Arginine, Histidine

Structures and classifications of amino acids

Polypeptide Chain Formation

Amino acids are joined by peptide bonds, forming linear polypeptide chains. Each chain has an amino (N) terminus and a carboxyl (C) terminus, defining the directionality of the protein.

Peptide bond: Covalent bond between the carboxyl group of one amino acid and the amino group of the next.
Directionality: Proteins are synthesized from N-terminus to C-terminus.

Protein Structure and Function

The three-dimensional structure of a protein determines its function. Proteins fold based on the chemical properties of their amino acids, and the forces stabilizing their conformation are weaker than covalent bonds.

Primary structure: Sequence of amino acids.
Secondary/tertiary/quaternary structure: Higher-order folding and assembly.
Function: Enzymatic activity, structural support, signaling, etc.

Protein 3D structure

Genetic Mutations and Disease: Sickle Cell Anemia

A single nucleotide change in DNA can alter the amino acid sequence of a protein, potentially causing disease. Sickle cell anemia is caused by a single amino acid substitution (Glu to Val) in the β-globin protein, affecting its structure and function.

Mutation: Point mutation in the β-globin gene.
Effect: Alters hemoglobin properties, leading to sickle-shaped red blood cells.

Amino acid change in sickle cell anemia Protein electrophoresis gel for sickle cell hemoglobin Comparison of normal and sickle cell hemoglobin structure and function

The Genetic Code

Triplet Nature of the Genetic Code

The genetic code is read in groups of three bases (codons), each specifying one amino acid. Experiments by Crick and Brenner using bacteriophage T4 mutants demonstrated the triplet nature of the code.

Codon: Three-base sequence in mRNA specifying an amino acid.
Triplet code: 43 = 64 possible codons, sufficient for 20 amino acids.

Bacteriophage infecting E. coli Suppression of mutations demonstrates triplet code Table of mutation combinations and phenotypes Genetic code table

Deciphering the Genetic Code

Marshall Nirenberg and Gobind Khorana used synthetic RNAs and cell-free extracts to determine which codons specify which amino acids. Their experiments established the complete genetic code.

PolyU experiment: UUU codes for phenylalanine.
PolyA experiment: AAA codes for lysine.
PolyC experiment: CCC codes for proline.
Repeating sequences: Used to identify codons for other amino acids.

Table of polypeptide production from synthetic mRNAs

Properties of the Genetic Code

The genetic code has several key properties:

Triplet: Each codon is three bases.
Non-overlapping: Codons are read one after another, not overlapping.
No punctuation: The code is read continuously.
Start and stop codons: AUG (start), UAG/UAA/UGA (stop).
Degenerate: Most amino acids are encoded by more than one codon.
Nearly universal: The code is conserved across most organisms.

Overlapping vs non-overlapping genetic code

Translation: Mechanism and Machinery

Ribosome Structure and Function

The ribosome is the molecular machine responsible for translation. It reads the mRNA sequence and assembles the corresponding amino acid chain.

Ribosome: Composed of rRNA and proteins, with large and small subunits.
Active sites: A (aminoacyl), P (peptidyl), and E (exit) sites for tRNA binding.

Ribosome structure and tRNA binding sites Ribosome structure in prokaryotes and eukaryotes

Translation in Prokaryotes vs Eukaryotes

Translation occurs differently in prokaryotes and eukaryotes. In prokaryotes, transcription and translation are coupled, while in eukaryotes, they are separated by the nuclear membrane.

Prokaryotes: Translation begins before transcription is complete.
Eukaryotes: mRNA is processed in the nucleus, then translated in the cytoplasm.

Coupled transcription and translation in bacteria

Types of RNA in Translation

Several types of RNA are involved in translation:

mRNA: Messenger RNA, carries the genetic code.
tRNA: Transfer RNA, brings amino acids to the ribosome.
rRNA: Ribosomal RNA, forms the core of the ribosome and catalyzes peptide bond formation.
miRNA/siRNA: Regulate gene expression.

Ribosome structure and tRNA binding sites Ribosome structure in prokaryotes and eukaryotes

tRNA Structure and Function

tRNAs are adapter molecules that recognize specific codons in mRNA and deliver the corresponding amino acid. Each tRNA has an anticodon loop and an amino acid attachment site.

Anticodon: Three-base sequence complementary to the mRNA codon.
Charging: Aminoacyl-tRNA synthetase enzymes attach the correct amino acid to the tRNA.

Aminoacyl-tRNA synthetase charging tRNA

Wobble Hypothesis

The third base of the codon (wobble position) allows for flexible pairing, enabling one tRNA to recognize multiple codons for the same amino acid.

Stages of Translation

Translation occurs in three stages: initiation, elongation, and termination.

Initiation: Ribosome assembles at the start codon.
Elongation: Amino acids are added one by one to the growing polypeptide chain.
Termination: Translation ends at a stop codon, and the polypeptide is released.

Translation initiation in E. coli Translation elongation: tRNA binding to A site Translation elongation: peptide bond formation Peptidyl transferase activity of ribosome Translation elongation: translocation Translation termination: stop codon recognition Translation termination: release factor action Ribosome recycling after translation

Summary Table: Properties of the Genetic Code

Property	Description
Triplet	Each codon is three bases
Non-overlapping	Codons are read one after another
No punctuation	Code is read continuously
Start/Stop codons	AUG (start), UAG/UAA/UGA (stop)
Degenerate	Most amino acids have multiple codons
Universal	Code is conserved across organisms

Key Equations

Number of possible codons:
Peptide bond formation:

Example: Sickle Cell Anemia

A single nucleotide change in the β-globin gene leads to a single amino acid substitution, which alters the protein's structure and function, causing sickle cell disease.

Additional info:

Some details about the triplet code and the experiments by Crick, Brenner, Nirenberg, and Khorana were inferred and expanded for academic completeness.