Protocol Online logo
Top : New Forum Archives (2009-): : Research Idea, Design and Collaboration

How do mutations distal to the active site effect activity? - (Aug/24/2011 )


I want to study how mutations distal to the active site effect the activity of the enzyme. I'm looking for a smaller protein 15KD to 35KD (I'll be using NMR and hi-throughput crystallization) which sees an increase in activity along a evolutionary path say E.coli -> Yeast, Yeast -> mammal. something within this span. I want a protein that depsite the increase in activity shows little change in the active site.

I know of a couple but all are too large.

any suggestions.

Thank you


1) One option is aaRS - aminoacyl-tRNA synthetases catalytic domains not very big and highly conserved. Some structures exist, some enzymes are bigger than others option to focus on editing or catalytic domains.
2) Another classic enzyme is from the glucose metabolism e.g. Glyceraldehyde-3-phosphate dehydrogenase A. There are crystal structures, on the upper limit of size for the NMR.

I think that is a very interesting subject. Highly relevant is almost any directed evolution enzyme for improved thermostability or processivity (often mutations are in the second or outer shell from the catalytic site).