Next Experiment: Western Blot of Myoglobin. Brief Overview Intro to Immunology. Figure 8-39b Representations of the X-ray structure of sperm whale myoglobin. ( b ) A diagram in which the protein is represented by its computer-generated C a backbone. Page 244.
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Western Blot of Myoglobin
Intro to Immunology
Figure 8-39bRepresentations of the X-ray structure of sperm whale myoglobin. (b)A diagram in which the protein is represented by its computer-generated Ca backbone.
Figure 8-39cRepresentations of the X-ray structure of sperm whale myoglobin. (c)A computer-generated cartoon drawing in an orientation similar to that of Part b.
Immobilized Metal Ion Affinity Chromatography (IMAC)
Immobilized Metal Ion Affinity Chromatography (IMAC) with Ni2+ is the standard method for purifying proteins with a lot of surface histidine or for engineered His-tagged proteins.
The (His)6-tag has become the most used affinity tag due to its small size, strong metal ion binding and ability to bind also under denaturing conditions.
How could you create a his tag?
In our experiment, the immobilized metal is Zn+2
Mb has 12 inherent his
with high Zn+2 affinity
How is this specificity mediated?
Complementarity: receptor/ligand binding is essentially pattern recognition between two complementary surfaces.
Ag: Angiotensin II
Ag: HIV protease fragment
Stryer Fig. 4.36 Western blot.
Antibodies make great
many Ab classes
one Ab class
Enzyme Linked Immunosorbent Assay (ELISA)