Use of exons in 2-Hybrid analysis

Dean Danner Lab djdlab at bimcore.emory.edu
Mon Jun 5 11:34:58 EST 1995


 I am currently using the two-hybrid system to look at protein-protein
interactions between known components of a multi-enzyme complex.  Up to
now, I have been using cloned DNA sequences which represent the full length
protein to fuse into the 2-hybrid vectors.  

Recently, my mentor told me that some groups have abandoned the use of
full length protein sequences in 2-hybrid analysis, using individual exons
instead.  I have several questions about this approach.

(1) What is the rationale behind using exons instead of a larger protein?
    It seems that isolation of individual component peptide sequences out of
    the "context" of the rest of the protein will increase the risk of non-
    specific interactions by charge or by hydrophobic interactions.
    Additionally, it seems that interactions which may require the presence 
    of multiple exons may not be uncovered by this method.

(2) If one is already able to measure interaction between two full length
     proteins using the 2-hybrid system, is there an advantage to using 
     the exon approach instead of deletion analysis of the full length 
     sequence?

(3)  What references and/or helpful hints can you give me on this method?

Thanking you in advance,
Brett Burkholder		Emory University
(Dean Danner lab)		Department of Genetics and Molecular Medicine
burkhold at gmm.gen.emory.edu









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