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How are nucleotides organized?
Codons � 3 letter nucleotides
Where does transcription take place?
Where does translation take place?
Cytoplasm � ribosomes
Eukayrotic RNA modification
- � 5� cap � 7-methyl guanosine cap
- o Help directs mRNA
- o Recruitment of ribosomes
- � 3� poly (A) tail
- o Multiple translations
What do transcriptions factors do?
Help orient RNA polymerase
7-methyl guanosine caps helps what?
Orientation in the ribosome
3� poly (A) tail helps so it can be used multiple times
RNA editing =
- Many products out of the same info
- Introduction of a stop cordon
- Differential splicing - cuts out introns and can put in different orientation to make a different product
Open reading frames
Sites where gene expression can begin
Furin � protein � enzyme � fes feps � upstream region fur
tRNA � enzymes to carry out this function
consensus sequences help orient enzymes
rRNA � catalytic activity
- acts as an enzyme
- splice out introns
RNA primers in eukaryotes are removed by
5� to 3� modifications prevents ribonuclease activity before it leave the nuclease.
The Central Dogma
The central dogma of biology is that information stored in DNA is transferred to RNA molecules during transcription and to proteins during translation.
Transcription and Translation in Prokaryotes
- � The primary transcript is equivalent to the mRNA molecule.
- � The mRNA codons on the mRNA are translated into an amino acid sequence by the ribosomes.
Transcription and Translation in Eukaryotes
- � The primary transcript (pre-mRNA) is a precursor to the mRNA.
- � The pre-mRNA is modified at both ends, and introns are removed to produce the mRNA.
- � After processing, the mRNA is exported to the cytoplasm for translation by ribosomes.
Messenger RNAs (mRNAs)�
intermediates that carry genetic information from DNA to the ribosomes.
Transfer RNAs (tRNAs)�
adaptors between amino acids and the codons in mRNA.
Ribosomal RNAs (rRNAs)�
structural and catalytic components of ribosomes.
Small nuclear RNAs (snRNAs)�
structural components of spliceosomes.
Micro RNAs (miRNAs)�
short single-stranded RNAs that block expression of complementary mRNAs.
The central dogma of molecular biology is that
genetic information flows from DNA to DNA during chromosome replication, from DNA to RNA during transcription, and from RNA to protein during translation.
Transcription involves the synthesis of an RNA transcript complementary to one strand of DNA of a gene.
Translation is the conversion of information stored in the sequence of nucleotides in the RNA transcript into the sequence of amino acids in the polypeptide gene product, according to the specifications of the genetic code.
The Process of Gene Expression
- Information stored in the nucleotide sequences of genes is translated into the amino acid sequences of proteins through unstable intermediaries called
- messenger RNAs.
Evidence for an Unstable Messenger RNA
- � Synthesis of viral proteins in infected bacteria involved an unstable RNA molecule synthesized from the viral DNA.
- � Phage proteins were synthesized by bacterial ribosomes.
RNA Synthesis And Transport in Eukaryotes
- � Method: Pulse-Chase Labeling
- � At first, labeled RNA is exclusively in the nucleus.
- � Later, the labeled RNA is found in the cytoplasm.
- � RNA is synthesized in the nucleus and then transported to the cytoplasm.
General Features of RNA Synthesis - Similar to DNA Synthesis except
- �The precursors are ribonucleoside triphosphates.
- �Only one strand of DNA is used as a template.
- �RNA chains can be initiated de novo(no primer required).
The RNA molecule will be complementary to the DNA template (antisense) strand and identical (except that uridine replaces thymidine) to the DNA nontemplate (sense) strand.
RNA synthesis is catalyzed by RNA polymerases and proceeds in the 5� to 3� direction.
In eukaryotes, genes are present in the nucleus, whereas polypeptides are synthesized in the
Messenger RNA molecules function as intermediaries that carry genetic information from DNA to the
ribosomes, where proteins are synthesized.
RNA synthesis, catalyzed by RNA polymerases, is similar to DNA synthesis in many respects.
RNA synthesis occurs within a localized region of strand separation, and only one strand of DNA functions as a template for RNA synthesis.
Transcription in Prokaryotes
Transcription�the first step in gene expression�transfers the genetic information stored in DNA�genes�into messenger RNA molecules that carry the information to the ribosomes�the sites of protein synthesis�in the cytoplasm.
E. Coli RNA Polymerase
- � Tetrameric core: alpha2, beta ?beta�
- � Holoenzyme: alpha2, beta. Beta�?, sigma
- � Functions of the subunits:
- � alpha: assembly of the tetrameric core
- � beta: ribonucleoside triphosphate binding site
- � beta�: DNA template binding region
- � sigma: initiation of transcription
Initiation of RNA Chains
- � 1.Binding of RNA polymerase holoenzyme to a promoter region in DNA
- � 2.Localized unwinding of the two strands of DNA by RNA polymerase to provide a single-stranded template
- � 3.Formation of phosphodiester bonds between the first few ribonucleotides in the anscent RNA chain
Numbering of a Transcription Unit
- � The transcript initiation site is +1.
- � Bases preceding the initiation site are given minus (�) prefixes and are referred to as upstream sequences.
- � Bases following the initiation site are given plus (+) prefixes and are referred to as downstream sequences.
A Typical E. coli Promoter
- � Consensus sequences: -10 sequence and -35 sequence
- � Recognition sequence: -35 sequence
Termination Signals in E. coli
- � ?Rho-dependent terminators�require a protein factor (?)
- � ?Rho-independent terminators�do not require ?
RNA synthesis occurs in three stages:
(1) initiation, (2) elongation, and (3) termination.
RNA polymerases�the enzymes that catalyze transcription�
are complex multimeric proteins.
The covalent extension of RNA chains occurs within locally unwound segments of
Chain elongation stops when RNA polymerase encounters a
Transcription, translocation, and degradation of mRNA molecules often occur simultaneously in prokaryotes.
Transcription and RNA Processing in Eukaryotes
Three different enzymes catalyze transcription in eukaryotes, and the resulting RNA transcripts undergo three important modifications, including the excision of noncoding sequences called introns. The nucleotide sequenced of some RNA transcripts are modified posttranscriptionally by RNA editing.
Modifications to Eukaryotic pre-mRNAs
- � A 7-Methyl guanosine cap is added to the 5� end of the primary transcript by a 5�-5� phosphate linkage.
- � A poly(A) tail (a 20-200 nucleotide polyadenosine tract) is added to the 3� end of the transcript. The 3� end is generated by cleavage rather than by termination.
- � When present, intron sequences are spliced out of the transcript.
- Usually the genetic information is not altered in the mRNA intermediary.
- Sometimes RNA editing changes the information content of genes by
- � �Changing the structures of individual bases
- � �Inserting or deleting uridine monophosphate residues.
Three different RNA polymerases are present in eukaryotes, and each polymerase transcribes a distinct set of genes.
Eukaryotic gene transcripts usually undergo three major modifications:
- (1) The addition of 7-methyl guanosine caps to t� termini,
- (2)The addition of poly(A) tails to 3� ends, and
- (3)The excision of noncoding intron sequences.
The information content of some eukaryotic transcripts is altered by RNA editing, which changes the nucleotide sequences of transcripts prior to their translation.
Interrupted Genes in Eukaryotes: Exons and Introns
Most eukaryotic genes contain noncoding sequences called introns that interrupt the coding sequences, or exons. The introns are excised from the RNA transcripts prior to their transport to the cytoplasm.
- � Introns (or intervening sequences) are noncoding sequences located between coding sequences.
- � Introns are removed from the pre-mRNA and are not present in the mRNA.
- � Exons (both coding and noncoding sequences) are composed of the sequences that remain in the mature mRNA after splicing.
- � Introns are variable in size and may be very large.
Most, but not all, eukaryotic genes are split into coding sequences called exons and noncoding sequences called introns.
Some genes contain very large introns; others harbor large number of small introns.
The biological significance of introns is still open to debate.
Removal of Intron Sequences by RNA Splicing
The noncoding introns are excised from gene transcripts by several different mechanisms.
- � Removal of introns must be very precise.
- � Conserved sequences for removal of the introns of nuclear mRNA genes are minimal.
- � �Dinucleotide sequences at the 5� and 3� ends of introns.
- � �TACTAAC box about 30 nucleotides upstream from the 3� splice site.
Types of Intron Excision
- � The introns of tRNA precursors are excised by precise endonucleolytic cleavage and ligation reactions catalyzed by special splicing endonuclease and ligase activities.
- � The introns of some rRNA precursors are removed autocatalytically in a unique reaction mediated by the RNA molecule itself.
- � The introns of nuclear pre-mRNA (hnRNA) transcripts are spliced out in two-step reactions carried out by spliceosomes.
tRNA Splicing in Yeast
- ?A splicing endonuclease makes two cuts at the end of the intron.
- ?A splicing ligase joins the two ends of the tRNA to produce the mature tRNA.
- ?Specificity resides in the three-dimensional structure of the tRNA precursor, not in the nucleotide sequence.
- � RNA/protein structure
- � Excises introns from nuclear pre-mRNA
Noncoding intron sequences are excised from RNA transcripts in the nucleus prior to the transport to the cytoplasm.
Introns in tRNA precursors are removed by the concerted action of a splicing endonuclease and ligase, whereas introns in some rRNA precursors are spliced out autocatalytically�with no catalytic protein involved.
The introns in nuclear pre-mRNAs are excised on complex ribonucleoprotein structures called spliceosomes.
The intron excision process must be precise, with accuracy to the nucleotide level, to ensure that codons in exons distal to introns are read correctly during translation.
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