Unit 3 notes from textbook + slides Flashcards
where does transcription and translation occur in bacteria?
cytoplasm
where does transcription and translation occur in eukaryotes?
transcription occurs in the nucleus and translation in the cytoplasm
exception to the central dogma
viruses are an exception and can go from RNA->DNA or RNA->RNA
difference between uracil and thymine
thymine has a methyl group on its 5’ carbon whereas uracil has a hydrogen
transcription definition
as a region of DNA unwinds, one strand is used as a template for the synthesis of an RNA transcript that is complementary in sequence
uses RNA polymerase
acts by adding nucleotides to the 3’ end of the growing transcript
stages of transcription
1) initiation: RNA polymerase and other proteins are attracted to double-stranded DNA, the DNA strands are seperated and transcription of the template strand begins
2) elongation: successive nucleotides are added to the 3’ end of the growing RNA transcript as the RNA polymerase proceeds along the template strand
3) termination: the RNA polymerase encounters a sequence in the template strand that causes transcription to stop and the RNA transcript to be released
directionality of the RNA transcript
synthesized in the 5’ to 3’ direction, the DNA template is read in the 3’ to 5’ direction
promoter definition
regions of typically a few hundred base pairs where RNA polymerase and associated proteins bind to DNA
Promoters in eukaryotes
includes the sequence 5’-TATAAA-3’ or something similar which is known as the TATA box
terminator sequence
transcription continues until the RNA polymerase encounters a sequence known as a terminator. transcription stops at the terminator and the transcript is released
how is promoter recognition mediated in bacteria?
a protein called sigma factor which associates with RNA polymerase and facilitates its binding to specific promoters
once transcription is initiated, sigma factor dissociates and the RNA polymerase continues transcription on its own
how is promoter recognition mediated in eukaryotes?
transcription in eukaryotes requires the combined action of at least 6 proteins known as general transcription factors that assemble at the promoter of a gene
also transcriptional activator proteins help control when and in which cells transcription of a gene will occur. they are able to bind with enhancer DNA sequences as well as with proteins that allow transcription to begin.
messenger RNA definition
RNA molecules that combines with ribosome to direct protein synthesis, it carries the genetic “message” from the DNA to the ribosome
RNA processing definition
converts the primary transcript into the finished mRNA which can then be translated by the ribosome
happens in the nucleus
RNA processing (3 types of chemical modifications)
1) the 5’ end of the primary transcript is modified by the addition of a special nucleotide. called the 5’ cap. the cap is linked to the RNA transcript by a triphosphate bridge between the 5’ carbons of both ribose sugars. the ribosome recognizes an mRNA by its 5’ cap
2) polyadenation: the addition of a string of about 250 consecutive A-bearing nucleotides to the 3’ end, forming a poly(A)tail. it stabilizes the RNA transcript until it is translated in the cytoplasm
3) not every stretch of the RNA transcript ends up being translated into protein. exons: regions of protein-coding sequence that are expressed introns: non-coding regions that are interspersed.
process of removal of the non coding introns and joining of the exons known as RNA splicing
ribosomal RNA
makes up the bulk of ribosomes and is essential in translation
transfer RNA
carries individual amino acids for use in translation
large and small subunit of the ribosome
the large subunit of the ribosome includes three binding sites for molecules of tRNA, which are called aminoacyl(A)site, peptidyl(P)site and the exit(E)site
use of ribosomes in translation
structures made up of RNA and protein that bind with mRNA and are at the site of translation
anticodon (tRNA)
three bases in the anticodon loop
the 3 nucleotides that undergo base pairing with the corresponding codon
anti parallel and complementary to the codon on the mRNA
aminoacyl tRNA synthetases
enzymes that connect specific amino acids to specific tRNA molecules
these enzymes are directly responsible for actually translating the codon sequence from a nucleic acid to a specific amino acid in a polypeptide chain
binds to multiple sites on any tRNA that has an anticodon corresponding to the amino acid and it catalyses formation of the covalent bond between the amino acid and tRNA
directionality of synthesis of polypeptide
from the amino end to the carboxyl end
translation stages
1) initiation: the intiator AUG codon is recognized and Met is established as the first amino acid of the new polypeptide chain
2) elongation: successive amino acids are added one by one to the growing chain
3) termination: addition of amino acids stops and the completed polypeptide chain is released from the ribosome
what does the initiation of translation require?
a number of protein intiation factors that bind to the mRNA. In eukaryotes, one group of intiation factors binds to the 5’ cap. Initiation factors recruit the small ribosomal subunit and tRNA met and scan the mRNA for an AUG codon. When the complex reaches an AUG the large ribosomal subunit joins and intiation factors are released.
translational elongation steps in depth
1) a tRNA complementary to the next codon binds to the A site
2) a reaction transfers the Met to the amino acid on the tRNA in the A site, forming a peptide bond
3) the ribosome moves down one codon which puts the tRNA carrying the polypetide into the P site and the now uncharged tRNA into the E site where it is released. A new tRNA complementary to the next codon binds to the A site
4) the polypeptide transfers to the amino acid on the tRNA in the A site. The polypeptide is elongated by repeating the previous two steps
what happens when the ribosome encounters a stop codon?
a protein release factor binds to its A site, causes the bond to break and ribosomal subunits dissociate
difference between bac and euk in terms of where the intiation complex forms
euk: forms at the 5’ end
bac: shine-dalgarno sequence
point mutation definition
where one base pair is replaced by a different base pair
silent mutation definition
a mutation where the change in a base still codes for the same amino acid
missense mutation definition
a mutation where the change in a base doesnt code for the same amino acid
nonsense mutation definition
creates a stop codon that prematurely terminates translation
deletion/insertion mutation definition
deletion or insertion of a small number of nucleotides, doesn’t affect non coding DNA
frameshift mutation definition
removing/inserting only one base, shifts the reading frame, the protein doesn’t fold properly and is not functional
positive regulation definition
a regulatory molecule (usually a protein) must bind to the DNA at a site near the gene in order for transcription to take place
negative regulation definition
a regulatory molecule must bind to the DNA at a site near the gene in order for transcription to be prevented
transcriptional activator protein definition
regulatory protein, activates transcription, RNA polymerase is recruited to the promoter of the gene by the activator
inducer definition
small molecule that interacts with the repressor and prevents it from binding DNA and blocking transcription
polycistronic mRNAs
the ability to initiate translation internally allows prokaryotic mRNAs to code for more than one protein
translation in prokaryotes initiation
the ribosome binds at any shine dalgarno sequence, with translation beginning at the next downstream AUG codon. this allows a single mRNA to code for multiple polypeptides
lac operon
lacI encodes the repressor protein
lacZ (cleaves lactose into glucose and galactose) and lacY (transporter protein) encode proteins necessary for the breakdown of lactose
lacO operator, binding site for the repressor
RBS in operons
each of the coding sites is preceded by a ribosome-binding site, so translation can be intiated at each coding sequence
In the absence of lactose…
the repressor binds to the operator and prevents transcription from taking place
In the presence of lactose…
the repressor protein is unable to bind to the operator because allolactose binds to the repressor changing its shape, when the operator is not bound to the repressor the promoter recruits the RNA polymerase complex and transcription occurs
mutation in the repressor protein results in..
trasncription occuring constitutively
mutation in the operator region results in..
transcription occuring constitutively
bacterial promoters for transcription
the -10 box is found 10 bases upstream from the transcription start site (+1 site)
the -35 box is 35 bases upstream from the +1 site
sigma factor binds to regions
what causes the mRNA to dissociate from the gene in bacteria?
hairpin loop
DNA binding proteins definition
bind in the major and minor grooves of DNA
bind by H-bonds and non-covalent interactions
side chains of amino acids interact with specific bases
recognize specific sequences of base pairs
How can transcription of a gene be regulated or controlled?
1) how often RNA polymerase plus sigma factor or other transcription factors bind to the promoter
2) how tightly RNA polymerase (+sigma or other transcription factors) bind to the promoter
alternative splicing and splice variants definition
different splicing pattern, more than one type of protein can be made from a single gene coding for an mRNA
what catalyses the peptide bond?
rRNA
how does the ribosome recognize the correct tRNA?
it doesnt
charged tRNAs are floating around the cytosol and diffuse in and out of the A site
open reading frame definition
the sequence of bases from and including the start codon to the stop codon (including)
