138 Zero to Genetic Engineering Hero - Chapter 5 - Extracting your engineered proteins
After the start codon, the ribosome continues to move
along the coding region of the RNA transcript (Figure
5-18). Thousands of the 64 different tRNA molecules
complete the Four B’s of basic cell operation and,
when the correct tRNA enters a translating ribo-
some and its anticodon matches perfectly with the
RNA transcript codon present in the ribosome, the
ribosome takes and connects the amino acid to the
growing chain (Figure 5-16). As new tRNA come into
the ribosome, it releases the previous tRNA that lost
its amino acid to the growing protein chain.
As the ribosome is translating the amino acid chain,
the amino acids in the chain begin to interact with
one another. The 20 amino acids are each quite
unique - some are negatively charged, some posi-
tively charged, and some not charged (Figure 3-30).
Some of these have sizeable bulky side chains (argi-
nine), and some have small side chains (glycine). It
is this wide variety of amino acid side chains that
gives the amino acids different “personalities”. As
the amino acid chain gets longer during translation,
the different amino acids in the chain bump and bind
or repel each other to form a complex and beautiful
three-dimensional shape (Figure 3-28). This three-
dimensional shape of proteins is what results in their
function - like being colorful!
In Figure 5-19, you will find one product of trans-
lation, the crystal structure of a protein called Red
Fluorescent Protein (RFP). Just like the structure of
DNA you saw in Chapter 1 (Figure 1-19), you will see
the red, blue, and gray segments of this structure
which correspond to oxygen, nitrogen, and carbon,
In Figure 5-19 you will also see a “ribbon” throughout
the structure, and this is commonly used as a guide to
help visualize the intricate and beautiful “beta barrel”
structure of the protein but is not a physical part of
the protein. RFP is only one example of thousands
of proteins that E. coli make, all of which have differ-
ent 3D shapes. You’ll see that RFP is very small at 3.5
nanometers wide. Compared to a DNA helix at ~12
nanometers wide, proteins can be quite small.
Figure 5-19. A protein is a long chain of amino acids folded
up into a three-dimensional structure. In this example, red
uorescent protein (RFP) has a beta-barrel structure. RFP
is a small protein that is about 250 amino acids long and is
about 3.5 nanometers wide. Crystal structure data source:
RCSB PDB: 2VAD.
3.5 nanometers (0.0000000035 m)
Figure 5-18. RNA transcripts are transcribed by RNA poly-
merase and are made up of ribonucleotides. A transcript has
a ribosomal binding site (RBS), 2-6 nucleotide spacer, start
codon, protein coding region, and a stop codon.
Protein coding region
The methionine start codon Going Deeper 5-6
So does this mean that every protein starts with methionine? Yes, it does, at least when it is made initially.
E. coli cells have some protein enzymes called proteases that are able to clip off the ends of proteins after
translation. Typically, proteases will cut off several amino acids from either end of the protein, one of which
could be the starter methionine.
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