Ch_17
Chapter 17: Gene Expression: From Gene to Protein
Concept 17.1: Genes specify proteins via transcription and translation
The DNA inherited by an organism leads to specific traits by dictating the synthesis of proteins.
Proteins act as links between genotype (genetic makeup) and phenotype (physical appearance).
Gene expression consists of two primary stages:
Transcription: Synthesis of RNA using the information in DNA.
Translation: Synthesis of polypeptides using the information in mRNA.
The relationship between proteins and DNA was discovered through various experimental approaches.
Evidence from Studying Metabolic Defects
In 1902, British physician Archibald Garrod proposed that genes dictate phenotypes through enzymes that catalyze specific chemical reactions.
He theorized that symptoms of an inherited disease reflect an inability to synthesize a particular enzyme, leading to disruptions in metabolic pathways.
Nutritional Mutants in Neurospora: Scientific Inquiry (1 of 2)
George Beadle and Edward Tatum exposed Neurospora (bread mold) to X-rays, creating mutants that could not survive on minimal media.
Their colleagues Adrian Srb and Norman Horowitz identified three classes of arginine-deficient mutants.
Each mutant lacked a different enzyme necessary for synthesizing arginine.
Nutritional Mutants in Neurospora: Scientific Inquiry (2 of 2)
The experiments supported the one gene–one enzyme hypothesis, which states that the function of a gene is to dictate the production of a specific enzyme.
The Products of Gene Expression: A Developing Story
Not all proteins are enzymes, leading researchers to revise the hypothesis to one gene–one protein.
Many proteins consist of several polypeptides, each dictated by its own gene.
The hypothesis is now refined to one gene–one polypeptide; however, gene products are commonly referred to as proteins.
Basic Principles of Transcription and Translation (1 of 3)
RNA acts as a bridge between genes and protein synthesis.
Transcription occurs when RNA is synthesized using DNA information, producing messenger RNA (mRNA).
Translation is the process of synthesizing polypeptides from mRNA sequences.
Translation occurs at the ribosomes, which are the sites of protein synthesis.
Basic Principles of Transcription and Translation (2 of 3)
In prokaryotic organisms, translation of mRNA can start before transcription is complete.
In eukaryotic cells, the nuclear envelope separates transcription from translation.
Eukaryotic RNA transcripts undergo modifications through RNA processing to create the finished mRNA.
Basic Principles of Transcription and Translation (3 of 3)
The primary transcript is the initial RNA transcript from any gene before processing.
The central dogma describes the flow of genetic information:
ext{DNA}
ightarrow ext{RNA}
ightarrow ext{Protein}
The Genetic Code
The assembly instructions for proteins in DNA utilize a code consisting of 20 amino acids encoded by combinations of 4 nucleotide bases.
Question raised: How many nucleotides correspond to an amino acid?
The answer is that a triplet code is used, meaning each amino acid is coded by a set of three nucleotides (codon).
Codons: Triplets of Nucleotides (1 of 4)
The flow of information from gene to protein follows a triplet code, consisting of nonoverlapping three-nucleotide sequences.
These sequences are transcribed into complementary nonoverlapping three-nucleotide sequences on mRNA, which is then translated into amino acid chains to form polypeptides.
Codons: Triplets of Nucleotides (2 of 4)
One of the two DNA strands, known as the template strand, provides a blueprint for synthesizing the complementary mRNA transcript.
The template strand remains consistent in its role for a given gene, with the opposite strand potentially serving as the template for another gene further along the chromosome.
Codons: Triplets of Nucleotides (3 of 4)
Specific DNA sequences linked to the gene determine which strand serves as the template.
The mRNA molecule is complementary to the template strand, and during translation, codons are read in the 5′ → 3′ direction.
Codons: Triplets of Nucleotides (4 of 4)
The non-template strand is referred to as the coding strand, as its nucleotide sequence mirrors the codons (T in DNA replaces U in RNA).
Each codon corresponds to one of the 20 amino acids found within polypeptides.
Cracking the Code
By the mid-1960s, all 64 codons were deciphered.
Of these, 61 codons code for amino acids, while 3 serve as “stop” signals to terminate translation.
The genetic code exhibits redundancy, meaning multiple codons can correspond to a single amino acid, but it is not ambiguous; each codon corresponds to only one amino acid.
Codons must be interpreted in the correct reading frame, as incorrect groupings can lead to erroneous protein production.
Evolution of the Genetic Code
The genetic code is nearly universal, found in simple bacteria as well as complex animals.
This indicates a shared biological language likely present early in the evolution of life.
Concept 17.2: Transcription is the DNA-directed synthesis of RNA: A Closer Look
Transcription serves as the first stage of gene expression, wherein RNA polymerase catalyzes RNA synthesis, separating DNA strands and assembling RNA nucleotides.
RNA synthesis adheres to the same base-pairing principles as DNA, except uracil (U) substitutes for thymine (T).
Molecular Components of Transcription (1 of 2)
RNA polymerase is responsible for facilitating RNA synthesis by attaching to a sequence called the promoter.
The end of transcription in bacteria is marked by a sequence called the terminator.
The segment of DNA engaged in transcription is referred to as a transcription unit.
Molecular Components of Transcription (2 of 2)
The three stages of transcription include
Initiation: The process where RNA polymerase binds to DNA and begins RNA synthesis.
Elongation: RNA polymerase moves along the DNA, adding nucleotides to the growing RNA chain.
Termination: The sequence signaling the end of RNA transcription is reached.
RNA Polymerase Binding and Initiation of Transcription
Promoters indicate the transcription start point, usually extending several dozen nucleotides upstream.
In eukaryotic cells, proteins known as transcription factors assist RNA polymerase binding and initiate transcription.
The completed complex of transcription factors and RNA polymerase II forms the eukaryotic transcription initiation complex.
One essential element in eukaryotic promoters is the TATA box, which is a critical component for assembling the initiation complex.
Elongation of the RNA Strand
As RNA polymerase progresses along the DNA, it unwinds the double helix about 10–20 nucleotides at a time.
RNA nucleotides are added to the 3′ end of the growing RNA molecule, proceeding at an average rate of 40 nucleotides/second in eukaryotes.
Multiple RNA polymerases can transcribe the same gene simultaneously.
Termination of Transcription
The termination mechanisms differ between prokaryotic and eukaryotic organisms.
In bacteria, RNA polymerase stops at the terminator, allowing for immediate translation without further modification.
In contrast, eukaryotic RNA polymerase II transcribes the polyadenylation signal sequence, leading to RNA transcript release 10–35 nucleotides beyond this signal.
Concept 17.3: Eukaryotic cells modify RNA after transcription
In eukaryotic nuclei, enzymes modify pre-mRNA (RNA processing) prior to its export to the cytoplasm.
RNA processing entails alterations to both ends of the primary transcript and usually involves excising non-coding sections (introns) while splicing together the coding sections (exons).
Alteration of mRNA Ends
Each end of pre-mRNA undergoes specific modifications:
The 5′ end receives a modified nucleotide 5′ cap.
The 3′ end is equipped with a poly-A tail.
These modifications serve multiple functions:
Facilitate mRNA export to the cytoplasm.
Protect mRNA from degradation by hydrolytic enzymes.
Aid ribosomal attachment at the 5′ end, ensuring efficient translation.
Split Genes and RNA Splicing (1 of 2)
Most eukaryotic genes and their RNA transcripts contain long stretches of noncoding nucleotides known as introns, positioned between coding regions (exons).
RNA splicing removes introns, yielding the mature mRNA that will be translated into proteins.
Split Genes and RNA Splicing (2 of 2)
The removal of introns is facilitated by spliceosomes, complexes comprising a mix of proteins and small RNAs that recognize splice sites.
The spliceosome's RNA molecules also catalyze the splicing reaction, enabling precise exon inclusion in the final mRNA product.
Ribozymes
Ribozymes are catalytic RNA molecules that function as enzymes capable of splicing RNA.
Three characteristics allow RNA to catalyze reactions:
Formation of complex three-dimensional structures through self-pairing.
The presence of functional groups in some RNA bases that take part in catalysis.
The ability of RNA to bond with other nucleic acid molecules.
The Functional and Evolutionary Importance of Introns (1 of 2)
Certain introns contain sequences regulating gene expression and can significantly impact gene products.
Alternative RNA splicing can lead to one gene coding for multiple distinct polypeptides, broadening the potential protein diversity beyond the number of genes present in the genome.
The Functional and Evolutionary Importance of Introns (2 of 2)
Proteins typically have a modular structure, comprising various domains.
Different exons often correspond to different functional domains within a protein, and exon shuffling may give rise to the evolution of new proteins by recombining exons from various genes.
Concept 17.4: Translation is the RNA-directed synthesis of a polypeptide: A Closer Look
Translation entails the transfer of genetic information from mRNA to proteins, leading to polypeptide formation.
Molecular Components of Translation
Transfer RNA (tRNA) plays a critical role in translating an mRNA message into protein.
tRNAs ferry amino acids to the growing polypeptide chain in ribosomes, facilitating translation's intricate biochemical processes.
The Structure and Function of Transfer RNA (1 of 4)
Each tRNA molecule translates a specific mRNA codon into its respective amino acid, bearing a precise amino acid on one end and an anticodon on the other that pairs with a complementary mRNA codon.
The Structure and Function of Transfer RNA (2 of 4)
A typical tRNA molecule is approximately 80 nucleotides long and takes on a cloverleaf shape due to base-pairing interactions.
The Structure and Function of Transfer RNA (3 of 4)
Due to hydrogen bonding, tRNA assumes a twisted, three-dimensional shape, becoming L-shaped, with both the 5′ and 3′ ends positioned near one end; the protruding 3′ end attaches the corresponding amino acid.
The Structure and Function of Transfer RNA (4 of 4)
Accurate tRNA translation necessitates two fundamental molecular recognition events:
First, a correct match between the tRNA and its amino acid, facilitated by the enzyme aminoacyl-tRNA synthetase.
Second, pairing between the tRNA anticodon and the mRNA codon.
The phenomenon of wobble allows the third base of a codon to vary and enables one tRNA to correspond to multiple codons.
The Structure and Function of Ribosomes (1 of 2)
Ribosomes are essential for coupling tRNA anticodons with mRNA codons during protein synthesis.
Eukaryotic ribosomes are larger than bacterial ones and differ in molecular composition.
Some antibiotics specifically inhibit bacterial ribosomes without affecting eukaryotic ribosomes.
Ribosomes consist of large and small subunits made from proteins and ribosomal RNAs (rRNAs).
The Structure and Function of Ribosomes (2 of 2)
A ribosome is equipped with three tRNA binding sites:
P site: Holds the tRNA carrying the growing polypeptide chain.
A site: Holds the tRNA for the next amino acid to be added.
E site: Exit site where discharged tRNAs leave the ribosome.
Building a Polypeptide
Three phases characterize translation:
Initiation
Elongation
Termination
Each stage necessitates protein factors assisting in the translation process, and energy is needed for certain steps.
Ribosome Association and Initiation of Translation
The initiation of translation commences with the small ribosomal subunit binding to mRNA and an initiator tRNA (carrying methionine).
The subunit progresses along the mRNA until it identifies the start codon (AUG).
Proteins called initiation factors facilitate the large ribosomal subunit's arrival, completing the translation initiation complex.
Elongation of the Polypeptide Chain (1 of 2)
During elongation, amino acids are incrementally added to the C-terminus of the growing chain, with the process requiring elongation factors.
Elongation encompasses three steps:
Codon recognition
Peptide bond formation
Translocation
Energy expends in the first and third steps.
Elongation of the Polypeptide Chain (2 of 2)
Translation advances along the mRNA in a 5′ → 3′ direction, while the ribosome and mRNA move in relation to one another codon by codon.
The elongation cycle takes less than a tenth of a second in bacteria, with empty tRNAs departing from the E site, returning to the cytosol for reloading.
Termination of Translation
Elongation continues until a stop codon reaches the A site, prompting the acceptance of a release factor.
The release factor stimulates the addition of a water molecule instead of an amino acid, which leads to the cleavage of the polypeptide from the tRNA.
Following this reaction, the translation assembly disaggregates.
Completing and Targeting the Functional Protein
Translation alone may not suffice to produce a functional protein.
Post-translational modifications or targeting to specific cellular locations may be required for protein functionality.
Protein Folding and Post-Translational Modifications
A polypeptide chain naturally coils and folds into a specific three-dimensional shape during synthesis, leading to secondary and tertiary structures.
The primary structure of a protein correlates with its overall shape, which determines its biological function.
Before proteins can perform their specific roles, post-translational modifications may be necessary.
Targeting Polypeptides to Specific Locations (1 of 3)
Cells contain two types of ribosomes: free ribosomes in the cytosol and bound ribosomes attached to the endoplasmic reticulum (ER).
Free ribosomes primarily synthesize proteins for cytosolic function, whereas bound ribosomes produce proteins for the endomembrane system or secretion.
Ribosomes remain identical and can interchangeably function as free or bound based on the location of synthesis.
Targeting Polypeptides to Specific Locations (2 of 3)
Polypeptide synthesis begins in the cytosol and typically concludes there unless directed to attach to the ER by a signal peptide—a sequence of approximately 20 amino acids at the leading end of the polypeptide.
Targeting Polypeptides to Specific Locations (3 of 3)
A signal-recognition particle (SRP) binds to the signal peptide, guiding the ribosome to a receptor protein embedded in the ER membrane.
The signal peptide is enzymatically cleaved upon membrane entry, while diverse signal peptides target proteins to different organelles.
Making Multiple Polypeptides in Bacteria and Eukaryotes (1 of 3)
Polyribosomes (polysomes) allow multiple ribosomes to translate a single mRNA concurrently, enabling rapid polypeptide production.
Making Multiple Polypeptides in Bacteria and Eukaryotes (2 of 3)
In bacterial cells, transcription and translation are coupled, allowing newly synthesized proteins to promptly diffuse to their functional sites.
Making Multiple Polypeptides in Bacteria and Eukaryotes (3 of 3)
In eukaryotes, the nuclear envelope separates transcription from translation, and RNA undergoes processing prior to exiting the nucleus.
Concept 17.5: Mutations of One or a Few Nucleotides Can Affect Protein Structure and Function (1 of 2)
Mutations entail alterations in the genetic information of a cell.
Point mutations refer to changes affecting just one nucleotide pair within a gene.
A single nucleotide alteration in a DNA template can result in an aberrant protein, significantly impacting an organism's phenotype.