I am calculating the structure of the protein-protein complex. Binding interface of protein complex is validating by mutagenesis experiments. We constructed a triple mutant protein which hydrophobic residues are mutated to Glycine. One hydrophobic residue is located at a flexible loop region and other two on helix. These three hydrophobic residues are highly solvent exposed residues.
But most of the publications show that interface residues of protein complexes are mutated to Alanine for further validation of protein complex ?
Hi Srinivas,
Well, this is a complex questions, what you can count as a good test mutation in each particular case. In general it is better to avoid Gly due to its high permitted ramachadran angles. This makes it in general secondary-structure disruptive aa. Ala substitution therefore is much better for mutagenesis. You probably still can get away with Gly if you demonstrate identical CD spectra for mutant and wt proteins.
Thanks Oleksiy Kovtun !
Yes ! We can check by CD spectra of Mutants and It is reported that Glycine Mutation can induce conformation flexibility , Ala Mutation is Best Option . As per your suggestion I ll check CD spectra for Mutants .
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