Question

IL2-Receptor (IL-2R) is a receptor of IL-2 that is expressed in lymphocytes. Below is the crystal structure of IL-2 and IL-2R bound.

(A) Describe the different noncovalent bonds that exist in the graphic. Be sure to describe in terms of both functional groups and amino acids.

(B) Describe how you would characterize the binding of the two proteins.(Remember that multiple techniques are commonly used as validation).

(C) List at least five buffers that would be necessary to test when characterizing the binding in part B and why.

IL-2 Rα E1 D2 H120 M25 K35 T41 T37 Helix A 42 Y43 NS7 E68 D1 K64 R36 Y4 Helix B' IL2 E61 IL-2

Answer 1

Hi,

In a given graphic, There are two kind of non covalent interaction were found (from original article http://www.pnas.org/content/103/8/2788.full?tab=ds#T3)

A)

1, Hydrophobic interatcions in two D2, D2, domains

D1 = Phe-42^IL-2, Phe-44^IL-2, Tyr-45^IL-2, Pro-65^IL-2, and Leu-72^IL-2 (in IL2 domain)

D2 = Leu-2^α, Met-25^α, Leu-42^α, and Tyr-43^α (in IL-2Rα Domain)

2, Hydrophilic Interactions/ polar periphery

five ion pairs (Lys-38^α/Glu-61^IL-2, Arg-36^α/Glu-62^IL-2, Glu-1^α/Lys-35^IL-2, Asp-6^α/Arg-38^IL-2, and Glu-29^α/Lys-43^IL-2

B)

There are list of assays to fond out protein-protein interatcions/binding. viz( Native-PAGE, others like Co-Immunoprecipitation (co-IP), Pull-Down Assay, Crosslinking Protein Interaction Analysis, Label Transfer Protein Interaction Analysis,Far-Western Blot Analysis ).

In the Native PAGE shift assay, we will find the difference in mobilty of indivdual proteins and mixture of prieteins. i,e,. If two proteins are binding and they migrate in diffent spead relative to individual proteins. from which it can be concluded for their binding/interaction.

In the present case the native PAGE was used to analyze and confirmed by SDS/PAGE and MALDI-TOF spectrometry.

C)