You are new to the lab and your job is to test a new antibody made against the l
ID: 53148 • Letter: Y
Question
You are new to the lab and your job is to test a new antibody made against the lab’s favorite protein,
guerkin. You run a series of Western blots using cellular extracts from a variety of different mouse
tissues to see if the antibody recognizes the 50 kD guerkin protein. In almost all of the extracts, you
observe a strong band at 50 kD. However, in extracts from skeletal muscle, you always see a 70 kD
band. You’re pretty sure this band is specific to the guerkin protein because you don’t observe it in
skeletal muscle samples from mice lacking the guerkin gene. Your advisor thinks this is really interesting
and wants you to look at the genomic structure of the guerkin gene to see if you can find an explanation
for this observation.
Genomic organization of guerkin
The open boxes indicate exons and the filled boxes are introns. Nucleotide number is shown.
--- --- -------- ----- -------- -------
134 202 1052 1985 2536 3000 3546
A) Provide an explanation for the generation of 50 kD and 70 kD proteins that react with the anti-guerkin
antibodies. Be certain to illustrate the molecular differences between the two proteins. [Hint: the average
molecular weight of an amino acid is 110 Daltons] (6 points)
B) In what region of the two proteins would you predict the antibodies to bind in order to recognize both?
Explanation / Answer
Length of box
Number of nucleotides
Approximate number of amino acids
Box 1
202-134
68
22.6
Box 2
1052 – 202
850
283.3
Box 3
1985 – 1052
933
311
Box 4
2536 – 1985
551
183.6
Box 5
3000 – 2536
464
154.5
Box 6
3546 – 3000
546
182
a. The exact number of amino acids encoded by each of the exons might slightly vary depending on the exact molecular weight of amino acids present in the protein. Here we consider that the average molecular weight of an amino acid is 110 Daltons.
Approximate number of amino acids in 50 kD protein = 50000/110= 454.5 amino acids (~ Box 3 + Box 5)
Approximate number of amino acids in 70 kD protein = 70000/110= 636.36 amino acids (~ Box 3 + Box 5 + Box 6)
b. By analysing the above data we can predict that the antibodies for guerkin protein bind to Box 3 and Box 5 in order to recognize both the bands.
Length of box
Number of nucleotides
Approximate number of amino acids
Box 1
202-134
68
22.6
Box 2
1052 – 202
850
283.3
Box 3
1985 – 1052
933
311
Box 4
2536 – 1985
551
183.6
Box 5
3000 – 2536
464
154.5
Box 6
3546 – 3000
546
182
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