CBG exam 2 flashcards

Gene Expression: Gene to Protein

DNA replication happens during the S phase

Eukaryotes: animals, plants, fungi, protist

Prokaryotes: bacteria

·      We start off as one cell; a zygote; after the sperm fertilizes the egg

·      ~40 trillion cells are in a fully mature adult

o   Numerous cells must divide and differentiate to create a fully functioning organism

Central Dogma

DNA is transcribed to RNA and translated to protein

DNA to RNA = transcription

RNA to protein = translation

RNA to DNA = reverse transcription

Central Dogma control means endless possibilies?

How? Cut out a certain part of s gene and the cells only from the human ear and inject them into the rat

Why? To know how to grow a new ear

Gene expression: from template to product

Several types of RNA are required for regulation

·      mRNA (Messenger RNA)- carries the instructions from DNA to the ribosome to make protein.

·      rRNA (Ribosomal RNA)- Structural component of ribosomes, needed to form peptide bonds.

·      tRNA (Transfer RNA)- Aligns amino acid with proper codon.

·      snRNA (small nuclear RNA)- Required for splicing.

Key components of transcription

·      Promoter- Required to recruit RNA initiation complex.

·      RNA initiation complex- Needed for RNA polymerase function, one member is a helicase. TATA binding protein required to bind the promoter.

·      RNA Polymerase- Recruits nucleotides to make RNA.

Promoter

·      In eukaryotes the primary promoter is the TATA box (consensus sequence).

Promoter: DNA sequence to which RNA polymerase binds (initiation complex binds first). The region upstream of the transcription initiation site has 2 different consensus 6 base pair sequences. One at –10 and one at –35.

Transcription Overview

1.     RNA initiation complex bind TATA box.

a.     Specifically the first is TFIID (which contains TATA binding protein (TBP)).

2.     Then RNA polymerase binds to complex.

3.     Lastly remaining members of complex bind,

a.     One member being a helicase.

Parts of Transcription

·      Initiation- After all members of complex are present RNA polymerase starts recruiting nucleotides

·      Elongation-RNA polymerase moves 5’ to 3’ recruiting nucleotides.

·      Termination-RNA polymerase reaches termination sequence and then detaches.

Elongation

•       RNA polymerase recruit's uracil instead of thymine. RNA polymerase lacks proofreading activity. How can RNA polymerase afford to lack proofreading activity?

RNA processing and turnover

·      Transcriptional regulation is the major method to regulate gene expression, but RNA processing and turnover are also important

·      Pre RNAs are modified before transport to the cytoplasm. This includes modification of both ends and removal of introns

·      7-methylguanosine cap: a cap is placed on the 5’ end co-transcriptionally. It stabilizes the RNA and facilitates binding of RNA to the ribosome

·      Polyadenylation of tail: the hexanucleotide AAUAAA signals polyadenylation. A nuclease cleaves the RNA and a polyA polymerase adds a 200 A tail. This aids stability and translation into protein

·      Splicing: introns are removed by splicing from exons

RNA processing and turnover

The spliceosome mediates splicing

Spliceosomes: large complexes of protein and RNA that mediate the splicing reaction

Ribozymes: RNA molecules that catalyze cleavage of RNAs at a specific recognition sequence.

Transfer RNAs are needed for translation

·      Transfer RNAs (tRNAs): adaptors to align the correct amino acid (AA) on the right codon. Codons are 3 bases that code 1 AA. tRNAs are 70 – 80 nucleotides long with a clover leaf structure

·      Two major sites: an AA attachment site and an anticodon loop located at the opposite end to bind the right codon

·      Aminoacyl tRNA synthetases: enzymes that attach specific AA's to specific tRNAs

Ribosomes are needed for translation

Ribosomes : catalyze protein synthesis.  Two subunits composed of protein and RNA. Abundant (1x10⁶ in eukaryotic cells)

Ribosomal RNA deficiency disease

·      Treacher Collins Syndrome- Mutated TCO1 gene which is required for rRNA transcription.

Initiation, elongation, and termination

Initiation is complex in eukaryotes (>12 initiation factors) but simpler in prokaryotes. Starts when tRNA-methionine and mRNA bind to the small ribosomal subunit

The peptide chain elongates by subsequent addition of AAs as the ribosome moves down the mRNA. When a stop codon is reached (UAA), the reaction terminates, the protein is released and the ribosome dissociates from mRNA.

Codon Table

Read the codon table

Initiation of translation

Methionine encoded by AUG, initiates translation in both prokaryotes and eukaryotes

The 7- methyl guanosine cap of eukaryotic mRNA binds the ribosome and helps to align it for translation

Initiation, elongation and termination factors

The first step in prokaryotes is binding of 3 initiation factors to the ribosome. The mRNA and tRNA join, GTP is hydrolyzed and the 70S initiation complex is formed

Initiation in eukaryotic cells is complex and requires at least 12 initiation factors.  After initiation, elongation and termination factors are required for translation to be completed

Elongation of the peptide chain

Start with first amino acid, binding site, found in bacteria,

first amino acid comes to the P site

Everything else comes to the A

Peptide bond forms through dehydration reaction

Entire ribosome subunit shifts which pushed amino acid

First amino acid pops off and are then in the chain?

To create long polypeptide chain

Continues until release factor binds to ?

Termination of translation

The release factor breaks the bond between the peptide chain and the tRNA

The nascent protein, tRNA, and ribosome all dissociate

Polysomes: mRNAs are translated by multiple ribosomes at one time and are spaced about 200 bases apart

Classes of Mutations

Two classes of mutations:

Point mutations- A substitution mutation in which one base is switched for another base.

Three types of point mutation:

1)    Silent mutation

2)    Missense mutation

3)    Non-sense mutation

Frameshift mutations: A mutation that alters the entire reading frame of a mRNA sequence.

Two types of frameshifts:

1)    Insertion mutation

2)    Deletion mutation

Silent Mutations       

Silent mutation- A mutation that does not alter the amino acid sequence of a protein.

Example- UUA change to UUG

Missense Mutations

Missense mutation- A mutation that switches one amino acid for another.

Does nothing for the ...

Nonsense Mutations

Nonsense mutation- A mutation that results in an early stop codon.

Frameshift Mutations

Mutation that alters the reading frame, nucleotides are read three nucleotides at a time.

Normal sequence

AUG-CAC-UUG

Insertion Example

AUG-UCA-CUU-G

Deletion Example

AUG-ACU-UG

Protein folding and processing

Chaperones: proteins that facilitate folding of other proteins. They act as catalysts; they are not part of the folded complex

They work by assisting other proteins to self-assemble, possibly by stabilizing the unfolded intermediates

For example, proper folding might require both the carboxy and amino terminus. Chaperones maintain the protein until it is completely translated and ready to properly fold

Protein cleavage

Cleavage of the polypeptide chain is important for maturation

Signal sequence: 20 AA at amino terminal that targets secreted proteins to the endoplasmic reticulum.  This sequence is hydrophobic and assists the protein through the membrane

Signal peptidase: cleaves signal sequence and allows protein to enter the ER.

The cell cycle

Organism

^

Cells  2 cells

^

Organelle

^

Macromolecules

^

Atom

S phase = DNA replication

Transcription / central dogma happens anytime

In a single human, we have enough DNA to stretch from the surface of the earth to the moon

Heredity

·      Offspring inherit chromosomes containing genetic material from parental generation.

·      Chromosomes- made of two sister chromatids.

·      Sister chromatids- made of chromatin.

·      Chromatin made of histones and DNA.

G1= linear chromosome

S = after it replicates, each half of the X = a sister chromatid but the whole X is a chromatid

G2 = X

X shape means that the chromosome has already divided

Chromosome pairs

·      In humans there are 22 pairs of autosomes and 1 pair of sex chromosomes. 

Y = male

Don’t have a Y = female

Chromosomes in Species

·      Chromosome number vary between species, the number of chromosomes within species remain consistent.

·      Genes are carried on chromosomes, what are genes?

•       Coding regions in DNA

The cell cycle has distinct phases

Stages of the Cell Cycle

·      Interphase

·      Mitosis (Diploid/somatic cells)

·      Cytokinesis

Þ   The length of the cell cycle varies

Þ   Most adult cells divide slowly (once every few days or weeks)

Þ   Cancer cells and some normal cells (bone marrow, stomach) divide constantly.

Þ   Embryo cells also divide fast during cleavage

Interphase

·      G1 phase- Primary cell growth (cell organelles duplicated).

·      S- DNA synthesis

·      G2- Secondary growth (centrosomes form) 

·      G0-Cell leaves the cell cycle and enters cell cycle arrest.

Centrosomes

·      Centrosomes are organelles that organize microtubules. Contains 2 centrioles which are small cylinder-shaped organelles that help form mitotic spindle.

Mitosis

1.     Prophase

2.     Prometaphase

3.     Metaphase

4.     Anaphase

5.     Telophase

1.     Prophase

·      Mitotic spindle forms, chromosomes condense, and nuclear envelope disappears.

2.     Prometaphase

·      Kinetochore forms at the centromere.

o   Centromere = Middle of the chromosome that connects the two arms of the chromosome

·      Some kinetochores attach to centromere while others interact with opposite poles of cell

3.     Metaphase

·      Centrosomes go to opposite poles of the cell. Chromosomes line up along metaphase plate.

o   Sister chromatids are identical / no difference

4.     Anaphase

·      Cohesion protein that holds sister chromatids together are degraded and sister chromatids are separated to opposite poles.

5.     Telophase

·      Nuclear envelope reappears, chromosomes become less condense and microtubules disassemble.

o   Start unwrapping

o   Centrosome breaks apart

Cytokinesis

·      Animal cell forms cleavage furrow (actin filaments).

·      Plant cell forms cell plate from cell wall (vesicles).

Mitosis Diagram