113Zero to Genetic Engineering Hero - Chapter 4 - Genetic Engineering Your E. coli Cells
Figure 4-29. As RNA polymerase travels downstream un-
zipping the double-stranded DNA, it “reads” the (-) template
strand. Free-oating, complimentary ribonucleotides (C-G or
U-A or T-A) will bind strongly to the nucleotide of the DNA’s
template strand within the polymerase. This triggers the poly-
merase to attach the ribonucleotide to the growing RNA strand.
If a nucleotide doesn’t bind strongly (for example a C-A), it will
bump out of the RNA polymerase. Eventually, the correct one
will enter and be added to the growing string of RNA.
A, U, C, G
How does the RNA polymerase know when to stop
transcribing a gene? In E. coli, RNA polymerase will
stop transcribing a DNA sequence in one of three ways:
The RNA polymerase ‘slips off’ the DNA: If you
look back to Chapter 1, where you learned about
the structure of double-stranded DNA and the
zippering that occurs between complementary
nucleotides, you’ll notice in Figure 1-17 that there
are a different number of bonds (dashed lines)
between a G-C pair compared to an A-T pair. A
‘G-C’ complement has three bonds that hold the
complementary nucleotides together. An ‘A-T’
pair, has only two bonds. More bonds mean stron-
ger interactions, which means that the bonding
strength between A-T is weaker than G-C.
When RNA polymerase is riding along transcribing
DNA, it uses the bonds between the transcribed
RNA and the DNA (-) template strand to hold itself
connected to the DNA. A long stretch of repeat T’s in
the DNA’s (+) leading strand (which correspond to A’s
in the (-) template strand) results in a string of U’s in
each of which also only has two
bonds. This results in weak interactions between
the RNA polymerase and the DNA, and often the
RNA polymerase simply slips off. You will often nd
stretches of T’s in DNA (U’s in RNA) at the end of a
gene. These are placed to cause the RNA polymerase
to fall off of the DNA and stop transcription
The RNA strand folds up, causing RNA polymerase
to fall off - a ‘terminator’:
What would happen
if ribonucleotides of an RNA string were able to
interact with other ribonucleotides in the same
string? You’ve seen that two different strands of
DNA can come together to form a double helix. Can
something similar happen with RNA? Yes!
Because RNA transcripts are quite exible, they
can flip and flop around, allowing nucleotides
of the RNA strand to come into contact with one
another. A string of RNA can interact with itself,
and similar rules apply: A binds to U, G binds to C.
When this happens, a ‘hairpin structure’ can form.
In the example in Figure 4-30, you’ll see what is
called the bacteriophage 82 late gene terminator.
Two regions of the RNA transcript complement
well enough to form what is called a stem and loop.
The overall structure is called a hairpin, and the
formation of the hairpin structure creates physical
stress between the RNA transcript in the RNA poly-
merase. This causes the RNA polymerase to fall off
of the DNA. In most cases, there is also a poly-uri-
dine, also called poly-U, segment (U-U-U-U-U...)
immediately following the stem. This results in
the RNA polymerase slipping off of the DNA, as you
saw in the rst example.
Figure 4-30. A hairpin is a structure where RNA folds upon
itself to create a structure that causes the RNA polymerase to
get jammed up and detach from the DNA. Another name for
this is a ‘terminator’.
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