MCAT Biochemistry Chapter 7

Messenger RNA (mRNA)

• Carries the information specifying the amino acid sequence of the protein to the ribosome

  • transcribed from template DNA strands and undergo post transcriptional modifications

• In eukaryotes is monocistronic ;codes for one specific protein

Transfer RNA (tNRA)

• Responsible for converting the language of nucleic acids to language of amino acids and peptides 

• Includes a folded strand ofRNA with a 3 nucleotide anticodon that codes for the appropriate codon on mRNA 

• Charged  or activated tRNA carry a specific amino acid for protein synthesis 

aminoacyl-tRNA synthetase

has different forms each for a specific amino acid it actiates 

• Requires two high-energy bonds from ATP, an energy rich bond

• Transfers activated amino acid to the 3’ end of the correct tRNA

• High energy of aminoacyl bond is used to create a peptide bond during translation

Ribosomal RNA (rRNA)

• Synthesized in the nucleolus and functions in assembling the polypeptide sequence for the overall protein 

  • Has four rRNA strands with genes originating in nucleolus

• Ribizoymes: enzymes of ribsoome; made of RNA molecules instead of peptides

Codons

• Basic unit that codes for an amino acid; 3 letter nucleotide sequence 

• 64 codons that code for 20 amino acids

• Codon of the mRNA is recognized antiparallel by a complementary anticodon on a tRNA

• AUG codes for start of translation 

• UAA, UGA, UAG, code for termination of translation 

Degenerancy and Wobble

• The genetic code is degenerate because many codons can code for many amino acids 

  Wobble position: describes the third codon position designed to protect against mutations; mutations that happen to this position are silent

  • When amino acids have multiple codons, the first two letters of the codons are the same 

Expressed Point Mutations

• point mutations that affect the primary amino acid sequence by incorrectly sequencing one of the first two bases of a codon 

  • Missense 

  • Nonsense 

Transcription

•  is necessary because DNA cannot leave the nucleus, as it will be degraded 

• Thus, it must convert the genetic code to RNA

Transcription Initiation and Elogation

1. Helicase and topoisomerase and other enzymes are key in unwinding and stabilizing unwound strand similar to initiation of DNA replication 

2. The antisense DNA strand is used as template and mRNA is synthesized antiparallel and complementary in the 5’ to 3’ direction

3. RNA is transcribed until a termination sequence is reached 

   1. RNA pol detaches 

   2. DNA rewinds 

   3. RNA produced is heterogenous nuclear hnRNA

      1. hnRNA is post transcriptionally modified into mRNA

Eukaryotic Transcription Initation

1.  RNA polymerase II binds at TATA box at the promoter region (approximately at the -25th base) to initiate transcription

   1. Transcription factors help it locate this region

   2. Does not proofread or use a primer dissimilar from DNA pol

   3. Bases are numbered from point of transcription 

      1.  first base transcribed is plus one

      2. Base directly to the left is negative one 

Posttranscriptional Processing

• Before hnRNA can leave nucleus must undergo 3 modification steps

  • Splicign

  • Addition of 5’ CAP

  • 3’ Poly A Tail

• UTRs exist at each ends of mRNA for start/stop codons signaling the where to end modifications

  • mRNA exits nucleus at nuclear pores

Splicing

posttanscriptional modification

• snRNPs (small nuclear ribonucleoproteins) recognize 5’ and 3’ sites of introns and cut them into a lariat shape that is eventually degraded

Addition of 5’ CAP

1. A 7-methylguanylate triphosphate cap is added during the process of transcription and is recognized by the ribosome

• modifcation is necessary before transcribed RNA leaves nucleus

3’ Poly A Tail

1. A long poly A (many adenine bases) tail that prevents the mRNA sequence from being degraded when it leaves the nucleus

   1. Tail being to degrade as soon as it leaves the nucleus

Alternative Splicing

•  describes how the primary transcript of hnRNA (premodified mRNA) may be spliced together in different ways to produce multiple variants of proteins encoded by the same original gene 

  • Allows organisms to make many more different proteins from from a limited number of genes 

  • adds to genetic diversity

Ribosome: Translational Structure

• Has three binding sites 

  • A site: binds aminoacyl 

  • P site: binds peptide 

  • E site: exit site 

initiation of Translation

1. In eukaryotes. the small ribosomal subunit binds to the 5’ cap (for prokaryotes this is the shine dalgarno sequence) 

2. Charged initiator ttRNA aligns its anticodon with the P site and binds the AUG start codon

   1. Large subunit then binds to small unit via initiation factors

Elongation (translation)

Three step cycle repeated for each amino acid added to the protein after the initiator methionine 

1. Ribosome moves in 5’ to 3’ direction synthsizing protein from its amino (N-) to its carboxyl terminus (-C)

   1. A site: holds the incoming aminoacyl-tRNA complex 

      1. Determined by the mRNA codon within the A site 

   2. P site: holds the tRNA that carries the growing polypeptide chain 

      1. Where first amino amino acid methionine (AUG) binds

      2. Peptide bond forms as as polypeptide passes from tRNA in P site to tRNA in A site via peptidyl transferase 

         1. GTP is used for energy during bond formation 

   3. E site: where the uncharged tRNA pauses before exiting the ribosome, unbinding the mRNA

2. Elongation factors: assist by locating and recruiting aminoacyl-tRNA along with GTP, while helping to remove GDP once the energy has been used 

   1. Some eukaryotic proteins contain signal sequences which designate a particular destination for a protein

Termination of Translation

1. Stop codons move into A site and and a protein called release factor (RF) binds to termination codon 

2. Water is added to end of polypeptide chain and termination factors hydrolyze the completed chain 

Post-translational Processing

• Final synthesis of protein is completed via folding 

• Chaperone proteins aide the folding of these finished polypeptide chains 

• Some proteins are cleaved or, in case of quaternary structure, subunits come together 

  • Biomolecules may be added

Post-Translational Processing: Biomolecular Addition

• Biomolecules may be added to a translated protein via

  • Phosphorylation : addition of phosphate groups 

  • Carboxylation : addition of carboxylic acids

  • Glycosylation: addition of oligosaccharides 

  • prenylation : addition of lipid groups

Operon

• A cluster of genes transcribed as a single mRNA 

  • controls gene expression in prokaryotes

• Ex: trp operon of E. coli. Have 5 genes that regulate the manufacturing of the enzyme that makes tryptophan

Jacob-Monod Model

• used to describe the structure and function of operons 

  • Says that operons contain

    • structural genes: code protein of interest

    • Operator site: non transcribable region that binds a repressor

    • Promoter site: provides place for RNA pol to bind 

    • Regulator gene: codes for repressor protein

• Says that operons can be repressible or inducible

Inducible Systems

system whrere repressor binds tightly to operator system preventing RNA pol from binding to structural gene

• Under negative control; binding reduces transcriptional activity

• Inducers bind repressors to permit transcriptional activity 

Lac Operon as an Inducible System

• Assisted by binding of catabolite activator protein (CAP)

• CAP acts as transcriptional activator when glucose levels are low

  • Signals production of lactose to use as alternative energy source for metabolism 

  • ￬glucose →cAMP to bind CAP→conformational change to CAP→CAP binds promoter→↑lactose

  • Example of positive control: binding of molecule increases transcription

Repressible Systems

• Allow constant production of a protein product 

• Repressor made by regulator gene is inactive till it binds a corepressor 

  • Complex binds operator site to reduce transcriptional activity 

• Negative feedback: final structural product acts as corepressor

• negative control = binding reduces transcription

• Ex. trp operon 

  • When tryptophan is high, it acts as corepressor binding to repressor protein and reducing transcription

Transcription Factors

• help control gene expression in eukaryotes

• Transcription activating proteins that search the DNA looking for specific DNA-binding motifs 

  • Are trans regulators: travel through cell to regulate gene 

  • Have 2 recognizable domains 

    • DNA Binding domain

    • Activation Domain

DNA Binding Domain

• binds to a specific nucleotide sequence in the promoter region or a DNA response element 

Activation Domain

allows for binding of several transcription factors and other important regulatory proteins

Enhancers

• A group of several response elements outside the promoter which allow for control of one gene’s expression by multiple signals; may be 1000 bps away

• Increases likelihood that gene will be amplified because of the variety of signals that can increase transcription levels

  • cis regulators: in same vicinity as gene they control

  • Signal molecules bind to specific receptors 

Gene Duplication

• Method of amplifying gene expression by duplicating genes on the same chromosome or replicating genes by unwinding with helicase and isolating a specific gene 

Regulation of Chromatin Structure

• Histone Acetylation

• Histone Deacetylation

• DNA Methylation

• there are more but this is what the chapter focused on; look up 7.6 or last pgs of chapter 7

Histone Acetlyation

•  use of histone acetylases to acetylate lysine residues found in the amino terminal tail regions of histone proteins 

  • Decreases positive charge on lysines and weakens interaction histone with DNA  allowing for easier access for transcriptional machinery 

  • Raises gene expression 

Histone Deacetylation

• uses histone deacetylases to remove acetyl groups and lower gene expression levels 

DNA Methylation

• Add methyl groups to cytosine and adenine nucleotides

•  Often linked with silencing of gene expression 

• Heterochromatin regions are more heavily methylated