Question
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1. Use the imaginary gel mages below to answer the questions below. Use the molecular weight standards (in KDo) to estimate sizes (do not graph). three gels were looded with the same three purified proteins B.cand D (loaded in lones B,cond b) Gel 1 was run under non-reducing conditions (SDS) Gel 2 was run under reducing conditions (sDs and 6el 3 was run native (assume this tells the molecular weight in the native state) Use these images to answer the questions below. Ge 3 A Gel 2 50,7 355 288 la. Using all three gel images if the bond in Gel1.Lane B is at 120 KDa, estimate the number and size of the subunits making up the native protein. lb. Using all three gel images, if the bands in Gel 2, Lane Core at 20 and 45 KDa, estimate the number and size of the subunits making up the native protein. 1c, using all three gel images, if the band in 6el 2.Lane D is at 30 KDa, estimate the number and size of the subunits making up the native protein. 1d. What type of bonds or forces hold the subunits together in the protein in Ione B? (circle one) disulfide hydrogen bonds probably both neither, there is one subunit le. What type of bonds or forces hold the subunits together in the protein lone D? one) in (circle hydrogen bonds probably both disulfide neither, there is one subunit lf. Define the protein in lone B using the following system: homodimer, heterodimer,homotrimer. etc. Protein B is a:
Explanation / Answer
Answer 1a. In Gel 2, lane B contains two bands which indicate presence of two sub units. The smaller subunit is equivalent to third band of marker (50.7 KDa). The larger subunit will be 120 – 51 = 69 KDa.
Answer 1b. A total of three subunits are present in the protein C. In lane C of gel 1, one band of 90KDa indicates that two subunits of 45 kDa are bonded together (homodimer). One subunit of 20 KDa makes the third subunit.
Answer 1c. Homodimer of 30KDa i.e. two subunits of 30 KDa making a total protein of 60 KDa.
Answer 1d. Disulfide bonds.
Only in the presence of B-mercaptoethanol, both subunits got separated clearly indicating the presence of disulfide bond.
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