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> >ZOOHCC - 501: Molecular Biology (Theory)
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> >Unit 3: Transcription and Regulatory RNAs > >
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>Prokaryotic Transcription >
>Although prokaryotes and eukaryotes perform basically the same
transcription process, there are important differences in eukaryotic
membrane-bound nuclei. For nuclear-bound genes, transcription occurs in the
nucleus and the mRNA transcript must be transported to the cytoplasm. In
prokaryotes, which lack membrane-bound nuclei and other organelles,
transcription takes place in the cytoplasm of the cell. >
>RNA Polymerase >
>RNA polymerase is an enzyme that produces mRNA molecules (just as DNA
polymerase produces new DNA molecules during DNA replication). Prokaryotes
use the same RNA polymerase to transcribe all genes. In E. coli, the
polymerase is composed of five polypeptide subunits. These subunits are
assembled each time a gene is transcribed and disassembled when
transcription is complete. Each subunit has a specific role (which you don't
have to remember). Polymerases composed of all five subunits are called
holoenzymes. >
>Protein involve >
>Prokaryotes use the same RNA polymerase to transcribe all genes. In E.
coli, the polymerase consists of five polypeptide subunits, two of which are
identical. Four of these subunits, called α, α, β and β', make up the core
polymerase enzyme. These subunits are assembled each time a gene is
transcribed and disassembled when transcription is complete. Each subunit
has its own role. Two α subunits are required for the polymerase to assemble
on DNA. The β subunit binds to ribonucleoside triphosphates and consequently
becomes part of the 'newly born' mRNA molecule. β' binds to the DNA
template. A fifth subunit, σ, is involved only in transcription initiation.
It confers transcription specificity so that the polymerase initiates mRNA
synthesis at a convenient initiation site. Without σ, the core enzyme
transcribes from random sites to generate mRNA molecules that specify
gibberish proteins. A polymerase composed of all five subunits is called a
holoenzyme (a holoenzyme is a biochemically active compound composed of an
enzyme and its coenzyme). >
>Initiation >
>Transcription in prokaryotes (and eukaryotes) requires partial unwinding of
the DNA double helix at the mRNA synthesis region. The area of unwinding
is called the transcription bubble. A DNA sequence to which a protein or
enzyme involved in transcription binds to initiate the process is called a
promoter. In most cases, promoters are upstream of the genes they control.
Specific sequences in promoters are very important as they determine whether
the corresponding gene is always transcribed, occasionally transcribed, or
seldom transcribed. The structure and function of prokaryotic promoters are
relatively simple (Figure 1). A critical sequence in prokaryotic promoters
is located 10 bases before the transcription start site (-10) and is
commonly referred to as the TATA box. >
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T >o initiate transcription, the RNA polymerase holoenzyme assembles at the
promoter. The dissociation of σ allows the core enzyme to follow the DNA
template and synthesize mRNA by adding RNA nucleotides following
base-pairing rules, similar to how new DNA molecules are generated during
DNA replication. Only one of the two strands of DNA is transcribed. The
transcribed DNA strand is called the template strand because it is the
template for mRNA production. The mRNA product is complementary to the
template strand and nearly identical to the other DNA strand, called the
non-template strand, except that RNA contains uracil (U) instead of thymine
(T) found in DNA. The points are different. Like DNA polymerases, RNA
polymerases add new nucleotides to the 3'-OH group of the previous
nucleotide. This means that the growing strand of mRNA is synthesized in the
5'-3' direction. Since DNA is antiparallel, this means that RNA polymerase
moves in the 3' to 5' direction along the template strand >
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>As the elongation progresses, the hydrogen bonds connecting the
complementary base pairs of the DNA double helix are broken, so the DNA is
continuously unwound in front of the core enzyme (Figure 2). DNA is unwound
behind the core enzyme while hydrogen bonds are reformed. The base pairs
between DNA and RNA are not stable enough to maintain the stability of the
mRNA synthetic components. Instead, RNA polymerase acts as a stable linker
between the DNA template and the newly formed RNA strand, ensuring that
elongation is not prematurely terminated. >
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Termination
> >When a gene is transcribed, RNA polymerase must be told to detach from
the DNA template and release the newly made mRNA.There are two types of
termination signals, depending on the gene being transcribed. One is
protein-based and the other is RNA-based. Both termination signals are
based on specific DNA sequences near the end of the gene that cause the
polymerase to release the mRNA. >
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>In prokaryotic cells, by the time transcription is finished, the processes
of transcription and translation both occur in the cytoplasm, so they can
occur simultaneously, so that transcription is already required to start
making copies of the encoded protein. used (Fig. 3). In contrast,
transcription and translation do not occur simultaneously in eukaryotic
cells, as transcription occurs within the cell nucleus and translation
occurs outside the cytoplasm. >