Ap Bio Ch. 17

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Gene expression

Back 1

process by which DNA directs protein synthesis

Transcription and translation

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Garrod

Back 2

1909 first to suggest that genes dictate phenotypes through enzymes that catalyze specific chemical reactions in the cell

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Beadle and Tatum

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studied molds to develop the one gene-one enzyme hypothesis

The function of a gene is to dictate the production of a specific enzyme

Showed how a combination of genetics and biochemistry could be used to work out the steps in a metabolic pathway

Scientists later figured out that not all proteins are enzymes

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RNA

Back 4

Ribose
Uracil instead of Thymine
Single stranded

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DNA

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Deoxyribose
Thymine instead of Uracil
Double helix

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Transcription

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Synthesis of RNA under the direction of DNA

Occurs in nucleus

DNA provides a template for the assimilation of a copy of mRNA (messenger RNA)

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Translation

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Synthesis of a polypeptide which occurs under the direction of mRNA

Occurs at ribosomes

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Reasons for using RNA to make proteins instead of DNA

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Using RNA as an intermediate step protects the DNA

Using RNA as an intermediate step allows more copies of a protein to be made simultaneously

Each RNA transcript can be translated repeatedly

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Differences in protein synthesis in prokaryotes and eukaryotes

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Prokaryotes

Since there’s no nucleus, the DNA is not separated from the ribosomes

Translation of an mRNA can occur while its transcription is still in progress

termination is different

Eukaryotes

Transcription and translation occurs separately

Nuclear envelope separates transcription and translation

termination is different

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Primary transcript

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the initial RNA transcript that is not translated into protein

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• Triplet code

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→ the genetic instructions for a polypeptide chain are written in the DNA as a series of nonoverlapping three nucleotide words

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• Template strand

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• Template strand→ the one DNA strand that is transcribed during transcription to make mRNA

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• Codons

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• Codons→ the mRNA base triplets that code for the 20 amino acids
o Usually written in the 5→3 direction

• You must read codons from 5→3

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• Marshall Nirenberg 1961

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→ discovered the first codon
o UUU specifies the amino acid phenylalanine

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start codon

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• Methionine→ start codon, AUG
o Sometimes methionine is cut out of the finished protein, but all proteins start with methionine

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• RNA polymerase

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o Separates the DNA strands
o Synthesizes the RNA base pairs to make the RNA
o Only works from 5→ 3
o Don’t need a primer to start working

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• Promoter

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→ DNA sequence where RNA polymerase attaches and initiates transcription → determines which of the two DNA strands will serve as a template

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• Terminator (in prokaryotes)

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• Terminator (in prokaryotes)→ sequence that signals the end of transcription

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• Transcription unit

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• Transcription unit→ stretch of DNA that is transcribed into an RNA molecule

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• RNA Polymerase II

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• RNA Polymerase II→ synthesizes mRNA in eukaryotes

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• RNA Polymerase I and III

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→ transcribe RNA molecules that are not translated into protein in eukaryotes

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• Three stages of transcription

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o Initiation
• RNA polymerase binds to the promoter, DNA strands unwind, polymerase initiates RNA synthesis
o Elongation
• Polymerase moves downstream, unwinds DNA, elongates the RNA transcript 5→3, DNA strands reform into double helix afterwards
o Termination
• RNA transcript is released and polymerase detaches from DNA

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• Transcription factors

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proteins that mediate the binding of RNA polymerase and the initiation of transcription
o Once certain transcription factors are attached to the promoter, RNA polymerase II binds to it

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• Transcription initiation complex

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→ the completed assembly of transcription factors and RNA polymerase II bound to the promoter

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• TATA box

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o Nucleotide sequence containing TATA
o Eukaryotic promoters commonly include a TATA box
o Transcription factors recognize the TATA box and bind to it

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• Prokaryotic termination

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o Transcription proceeds through a terminator sequence in the DNA
o Transcribed terminator (on the RNA) functions as the termination signal which causes the polymerase to detach and release the transcript

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• Eukaryotic termination

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o Pre-mRNA is cut from the growing RNA chain while RNA polymerase II continues to transcribe the DNA
o When the RNA polymerase transcribes a sequence on the DNA called the polyadenylation signal sequence AAUAAA, it causes the release of the pre-mRNA
o the RNA polymerase keeps transcribing DNA
o Transcription is terminated when the polymerase eventually falls off the DNA

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RNA processing

what happens before RNA leaves the nucleus

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RNA splicing occurs

• Pre-mRNA is modified before leaving the nucleus
o methyl guanine 5’ cap-→ a guanine nucleotide and 3 phosphates are attached to the 5’ end
o Poly-A tail→ 50-250 adenine nucleotides are added to the 3’ end

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• Purpose of methyl guanine 5’ cap and poly A tail

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o Facilitate the export of the mature mRNA out of the nucleus
o Help protect the mRNA from degradation by hydrolytic enzymes
o Help ribosomes attach to the 5’ end of the mRNA

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• RNA Splicing

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removal of a large portion of the RNA molecule that is initially synthesized before it leaves the nucleus
o Most eukaryotic genes and their RNA transcripts have long noncoding stretches of nucleotides that need to be cut out
o The sequence of DNA nucleotides that codes for a eukaryotic polypeptide is usually not continuous


when SNRPS cut out the introns

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• Introns

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• Introns→ regions in nucleic acid that are not expressed

• Introns are cut out of the mRNA before it leaves the nucleus

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• Exons

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• Exons→ regions in nucleic acid that are expressed

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• snRNPs

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o recognize the nucleotide sequence at each end of an intron and cut the intron out
o located in the nucleus
o composed of RNA and protein
o snRNA is the RNA in a snRNP particle

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• Spliceosome

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a complex of several different snRNPs joined with additional proteins
o Interacts with certain sites along an intron to release the intron and join together the two exons

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• Ribozymes

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• Ribozymes→ RNA molecules that function as enzymes

• In some organisms RNA splicing can occur without proteins or additional RNA moleucles→ the intron automatically cuts itself out
• Pre-rRNA removes its own introns
• The structure of some ribozymes allow them to function as enzymes
• Discovery of ribozymes showed that not all biological catalysts were proteins

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o Alternative RNA splicing

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• Some genes can give rise to more than one polypeptide
o Alternative RNA splicing→ you can make more than one polypeptide from the same gene depending on which segments are treated as exons during RNA splicing

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• Domains

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discrete structural and functional regions in a protein
o Different exons code for the different domains of a protein

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• tRNA

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→ transfer RNA
o interprets mRNA to make polypeptides
o transfers amino acids from the cytoplasm to the ribosome
o the cell keeps its cytoplasm stocked with all 20 amino acids for protein synthesis


• tRNA translates mRNA into amino acids

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• anticodon

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→ the nucleotide triplet attached to the end of the tRNA
o it pairs up with the complementary side of the mRNA

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• Aminoacyl-tRNA synthetase

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o Joins the correct tRNA with the correct amino acid
o The enzyme’s active site only fits a specific combination of tRNA and amino acid
o So there are 20 different synthetases- one for each amino acid
o The enzyme uses ATP to synthesize the tRNA with the amino acid

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• Wobble

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o Some tRNAs can bind to more than one codon
o The base U on a tRNA can bind with A – or G at the 3’ end
o Relaxation of the base pairing rules allows tRNAs to bind to more than one codon
o This explains why the synonymous codons for a given amino acid differ in their third base but not in their first two bases

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• EPA sites in a Ribosome

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o E site→ exit site
o P site→ site where the polypeptide chain elongates
• Peptidyl-tRNA binding site
o A site→ enter site
• Aminoacyl-tRNA binding site

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• rRNA

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ribosomal RNA makes up the large and small subunits of the ribosome
o subunits are made in the nucleolus of eukaryotes
• rRNA is the most abundant type of RNA
• research suggests that rRNA is responsible for structure and function of a ribosome, not the proteins that make up the ribosome
• a ribosome can be categorized as a big ribozyme

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• Initiation

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Proteins called initiation factors bring together mRNA, tRNA, and the two subunits of the ribosome
o The small subunit binds to the mRNA and the tRNA containing methionine
o The small subunit finds the start codon AUG
o The large subunit attaches to the start codon
o Initiation complex→ complex of large subunit, small subunit, tRNA, and mRNA
o Energy is spent in the form of GTP in this process

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• Elongation

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amino acids are added one by one to the preceding amino acid
o Elongation factors→ proteins that facilitate elongation
o Uses energy in the form of GTP
o tRNA brings amino acids to the ribosome and the polypeptide elongates

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• Termination of translation

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o Elongation continues until a stop codon is read
o Release factor→ protein that binds to the stop codon in the A site
• Causes the hydrolysis of the polypeptide chain which releases the polypeptide chain

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Polyribosomes

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• A single mRNA is used to make many copies of a polypeptide simultaneously because several ribosomes can translate the message from one mRNA at the same time
• More than one ribosome can attach to the same mRNA
• Polyribosomes/Polysomes→ when there’s more than one ribosome attached to an mRNA
o Multiple copies of the protein can be synthesized at once

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• How do ribosomes become free or bound?

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o All polypeptides begin synthesis in the cytosol until the growing polypeptide cues the ribosome to attach itself to the ER

o Signal peptide→ a protein that targets the ribosome to be attached to the ER

• Polypeptide proteins destined for the endomembrane system or for secretion are marked with a signal peptide

• Other signal peptides can target polypeptides to mitochondria, chloroplasts, other organelles→ but translation must finish before it is moved to other organelles

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o Signal Recognition Particle

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RNA complex that recognizes the signal peptide
• It brings the ribosome to the ER

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• siRNA and miRNA

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• siRNA and miRNA→ involved in regulation of gene expression

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• Small nucleolar RNA snoRNA

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aids in the processing of pre-rRNA transcripts for ribosome subunit formation in the nucleolus

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• Properties of RNA that allow it to preform so many different functions

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o RNA can hydrogen bond to other nucleic acid molecules
o RNA can assume 3D shapes by forming hydrogen bonds
o RNA has functional groups that allow it to act as a catalyst (like in ribozymes)

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• Mutations

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• Mutations→ changes in the genetic material of a cell

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• Point mutations

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• Point mutations→ chemical changes in just one base pair of a gene
o Leads to the production of an abnormal protein
o Two categories→ base pair substitutions and base pair insertions/deletions

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• Base pair substitution

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o Replacement of one nucleotide with another nucleotide
o Sometimes base pair substitutions are referred to as silent mutations b/c they have no effect on the encoded protein

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Missense mutations

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→ when the altered codon still functions but not as well

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Nonsense mutation

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→ when a point mutation changes a codon into a stop codon

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• Insertions and deletions

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o Additions or losses of nucleotides in a gene
o More disastrous than substitutions because it messes up all of the amino acids after the mutation

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o Frameshift mutation

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o Frameshift mutation→ when an insertion/deletion mutation messes up all of the other amino acids after the mutation

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• Mutagens

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• Mutagens→ physical and chemical agents that interact with DNA to cause mutations

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• Spontaneous mutations

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mutations that arise from errors occurring during DNA replication repair or recombination