determining oligomeric structure of a protein

Jesus M. Sanz sanz at
Sat Mar 5 07:02:07 EST 1994

In article <2l675s$mna at>,
rsaldanh at (Roland J Saldanha) wrote:

> I am working on the mitochondrial tyrosyl synthetase that functions as both a 
> synthetase and as a facilitator of group I intron splicing.  In characterizing 
> the protein we find that it elutes from gel filtration columns with an apparent
> molecular weight of a tetramer (monomer MW=72,000).  However the protein 
> crosslinks as a dimer with dimethyl suberimidate and glutaraldehyde.  This 
> would suggest a highly elongated structure that has an aberrant mobility 
> (reflecting the elongated size) in gel filtration.
> I would welcome suggestions for other methods for demonstrating the oligomeric 
> structure of the protein.  I am aware that sedimentation analysis is the best 
> method of approaching this problem but do not have access to an analytical 
> ultracentrifuge and would welcome collaborative offers from anyone who has a 
> machine.
> We would also like to try native page but the protein is very basic (computer 
> calculated pI of 10.3).  I would thus welcome suggestions for native gels of 
> basic proteins and suitable standards for basic proteins.

    You could also try another crosslinking reagent with a different 
specificity. For instance, water-soluble
carbodiimides are good for crosslinking between carboxylates, and may be
where dimethyl suberimidate and glutaraldehyde cannot work. You can find
a nice review about bifunctional reagents in Methods Enzymology (91), 580

Dr. Jesus M. Sanz                            Tf. (0223) 402146
Centre for Protein Engineering.              Fax.(0223) 402140
Hills Road,                      e-mail:sanz at
Cambridge CB2 2QH, U.K.     			    
            "Life is hard and happiness is ephemeral"

More information about the Proteins mailing list