Below is a section of DNA which codes for part of a protein. The bottom (blue) strand in this example is the template strand, which is also called the minus (-) strand,or the sense strand. It is this strand that serves as a template for the mRNA synthesis. The enzyme RNA polymerase sythesizes an mRNA in the 5' to 3' direction complementary to the template strand. The opposite DNA strand (red) is called the coding strand, the nontemplate strand, the plus (+) strand, or the antisense strand.
1 2 3 4 MUTATIONS: G A T T 5' C C C C T G G A A C A G T G G A C A G C A G G A 3' 3' G G G G A C C T T G T C A C C T G T C G T C C T 5'
a. Write the mRNA sequence.
5' CCC CUG GAA CAG UGG ACA GCA GGA 3'
b. Write the amino acid sequence.
pro-leu-glu-gln-trp-thr-ala-gly
c. Consider the mutations shown in the top line. Suppose the single mutation 1 (T - G) occurs in the 5' to 3' strand, with a concomitant change in the coding strand, write the amino acid sequence for the new protein. What might happen to the new protein if the original amino acid was buried inside the protein? If it was exposed on the surface?
mRNA second codon changes to CGG; amino acid change leu--> arg
Buried: replace nonpolar aa with +. Native protein greatly destabilized. Protein will not fold correctly. Function greatly impaired
Surface: Probably no great affect on structure. Replace a residue which doesn't like to be there with one that does. However, the function of the protein might be affected. If the Leu was part of a nonpolar binding pocket on the protein, a nonpolar ligand would not bind there any more.
d. Repeat (c), but with mutation 2 instead.
mRNA second codon changes to AAA; amino acid change Glu--> Lys
Buried: replace - aa with + aa. Native protein greatly destabilized. The initial glu would only be buried if were in an ion pair with a + residue. Hence, protein will not fold correctly. Function greatly impaired
Surface: Probably no great affect on structure. Replace a residue which likes to be there with one that also does. However, the function of the protein might be affected. If the Glu was part of a charged binding pocket on the protein, a positively charged ligand would not bind there any more.
e. Repeat (c), but with mutation 3 instead.
mRNA second codon changes to UAG; amino acid change Gln--> stop
Buried or surface: Form truncated protein. If it is very short compared to the original, protein nonfunctional. If it occurred late in the sequence, may have minor consequences. Most likely, you're dealing with a "dead" protein.
f. Repeat (c), but with mutation 4 instead.
mRNA second codon changes to GUA; amino acid change Gly--> Val. This is the mutation that occurs in sickle cell anemia.
Buried: replace smallest aa which often at tight bends in a folded protein, with a large nonpolar residue. The packing of the protein as it folds would be significantly altered, and the proteins usual structure destabilized. Structure and function likely to be significantly altered.
Surface: Probably some affect on structure. Replace a residue which is ambivalent to being on surface with one that doesn't like to be there. But not all nonpolar residues can be buried on folding. However, there will now be a nonpolar patch on the surface which might cause the protein to aggregate. This happens in sickle cell anemia.