Protein complexes can be identified by a method called tandem affinity purificat

Question

Protein complexes can be identified by a method called tandem affinity purification (TAP).

2) Five proteins (A, B, C, D, and E) were identified by TAP and the protein content of the complexes isolated when specific proteins were tagged is illustrated in Figure 1.

Figure1: The protein content of complexes purified using 4 distinct tagged proteins was resolved by 1D SDSPAGE and visualized by Coomassie blue staining (Lanes 1 to 4). The tagged proteins were A (lane 1), B (lane 2), D (lane 3) and E (lane 4). Horizontal discontinuous lines represent the electrophoretic mobility of proteins whose sizes are indicated on the left of the figure.

a) Identify the five proteins (A,B,C,D, & E) by carefully labeling, in each lane, the bands on Figure 1. When necessary, indicate with an asterisk (*) the tagged form of a protein. Note that only the five proteins listed above are represented in Fig. 1, and that the 5 untagged proteins differ from each other by at least 10 kDa. Explain your reasoning.

b) List the proteins that compose EACH identified complex. Briefly explain your reasoning.

c) If the TAP affinity purified complexes were submitted to native gel electrophoresis how many band(s) would you detect in lanes 1 and 2. Briefly explain.

Explanation / Answer

Identification of proteins (A, B, C, D and E) in the lanes

If a protein is tagged that will show little bit up in position in the gel because that will increase the molecular weight. But the molecular weight will be less on minus the MW of a tag.

So Lane 1 the 50Kd protein is A, the lane 2 the protein is in front of 30 Kd [similar], the lane 3rd has 90 Kd protein D and lane 4 has 60 Kd of E protein. So the C protein will be in mid of 70 and 80 Kd because it didn’t moved a little bit.

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