Protein Synthesis Gizmo – Jason Bao

2 Images from the Case Study:

Case Summary:
Explain how you found out that Lucy has ADA-SCID.
She had high deoxyadenosine levels and no HIV proteins present.
How did the stop codon mutation in Lucy's ADA gene stop her ADA protein from working?
It caused an early stoppage in the amino acid sequence, therefore impacting the shape and structure
of the ADA protein.
Your scan of Lucy's ADA gene showed a mutation in the active site. Why was this mutation not a
problem?
It was a silent mutation so the mutated codon still coded for the same amino acid.
Explain how stem cell therapy helped Lucy break down deoxyadenosine.
This therapy uses CRISPR technology to replace the incorrect DNA sequence with the correct
DNA sequence and multiplies this correct DNA by allowing these stem cells to multiply and
eventually be put back into the patient’s bone marrow.
Synopsis of Lucy’s Case – the experiments performed, and the conclusions they provided in
order to diagnose her condition
Overall, in this gizmo, a young girl named Lucy was experiencing a string of numerous
infections that indicated a problem with her immune system, which doctors immediately linked to
her abnormally low white blood cell count. Because of this, her immune system could not
adequately fight off infections, which led to Lucy’s current condition.
In response to her immunodeficiency, scientists were able to narrow down two causes of
her condition – HIV or ADA Severe Combined Immunodeficiency (ADA-SCID). However, after
an analysis of her blood, the scientists were able to determine that because of her low white blood
cell count, lack of HIV proteins, and high concentration of deoxyadenosine, Lucy must be
experiencing ADA-SCID. This is a condition that arises when the amino acid sequence of ADA is
altered in such a way that causes ADA to fold improperly, which causes it to form a different shape
and renders it unable to break down the toxin deoxyadenosine.
As such, with this new information, the scientists had to more thoroughly analyze Lucy’s
ADA protein, which revealed a normal range of gene length and mRNA length yet a much smaller
amino acid sequence than normal. This hinted at three possibilities: a mutation that changed an
amino acid in ADA’s active site (active site mutation), early stop codons that ended the amino acid
sequence early, and a tRNA mutation that carried the wrong amino acids to add to the polypeptide
chain. When given this information and extraneous information about the makeup of Lucy’s blood,
the scientists hypothesized that since Lucy had a much shorter amino acid sequence than the
normal range, she must have a stop codon mutation, because an early stop codon sequence in the
amino acid will result in a shorter protein and shorter amino acid sequence.
 To prove or reject this hypothesis, the scientists performed two experiments: a tRNA
analysis and an investigation of mRNA. After analyzing some of the tRNA molecules, the
scientists determined that each tRNA was carrying the correct amino acid for its anticodon and
thus all the tRNA were normal, eliminating the possibility of a tRNA mutation. Next, while
investigating mRNA, the scientists discovered two mutations in Lucy’s mRNA transcript as shown
below.

Since the mutation in the active site was a silent mutation (coded for the same amino acid), this
mutation did not affect the shape of the active site. However, the second mutation created a
premature stop in Lucy’s amino acid sequence, which shortened her enzyme from 363 to 283
amino acids, causing her ADA enzyme to malfunction. Because of this, the second experiment
was fundamental to understanding the cause of Lucy’s condition – an early stoppage in the amino
acid sequence.
Finally, to combat ADA-SCID, the scientists needed to replace this point substitution
nonsense mutation (UGA) with the normal codon (UCA) through the use of stem cell therapy.
Specifically, for Lucy, the scientists needed stem cells that could change into white blood cells
with working ADA enzymes. Thus, by using a technology called CRISPR/Cas9 that replaced the
incorrect DNA sequence of the ADA gene with the correct DNA sequence, the scientists enabled
these corrected stem cells to duplicate and eventually be injected into Lucy’s bone marrow,
eventually creating white blood cells with working ADA enzymes that break down the toxin,
strengthen Lucy’s immune system, and most importantly, cure her condition.