Hi Frank, I forgot to mention that the once purified, the protein is fairly stable and can be frozen and re-used for continued crystallization experiments. So, even though it is not available for purchase, it is a convenient system and having worked on it, it seems to me to be a model protein-DNA complex system almost equivalent to the lysozyme model. Thanks, Debanu.
-- Debanu Das, JCSG, SSRL. -----Original Message----- From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Das, Debanu Sent: Wednesday, August 15, 2007 11:31 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] model DNA-binding crystals? Hi Frank, The N-terminal catalytic fragment/polymerase domain of the MuLV Reverse Transcriptase is easy to express (large yield of soluble protein in E. coli expression, ~270 aa), purify (3 steps) and crystallize (overnight to couple of days with microseeding) with short stretches of DNA (8-16 bp) of different sequences to produce well diffracting protein-DNA complex crystals (1.5-2.5A) where the DNA structure is not influenced by lattice packing interactions as there are no direct crystal contacts between the DNA molecules. This particular system has been used to study the structures of some interesting DNA sequences as well as used in a kind of "host-guest" approach to study binding of DNA drugs to DNA. You can refer to the following references and the references therein: Cote M.L., Georgiadis M.M. Structure of a pseudo-16mer DNA with stacked guanines and two G-A mispairs complexed with the N-terminal fragment of Moloney murine leukemia virus reverse transcriptase. Acta Crystallogr. D Biol. Crystallogr. 2001;57:1238-1250 Goodwin, K. D., Long, E. C., and Georgiadis, M. M. (2005) A host-guest approach for determining drug-DNA interactions: An example using netropsin, Nucleic Acids Res. 33, 4106-4116. Regards, Debanu. -- Debanu Das, JCSG, SSRL. -----Original Message----- From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Frank von Delft Sent: Wednesday, August 15, 2007 11:14 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] model DNA-binding crystals? Hi, anyone know of a lysozyme-equivalents for protein-DNA complexes? i.e. a protein that a) is easy to obtain (preferably purchase) b) binds some bit of DNA well c) crystallizes easily as protein-DNA complex Cheers phx
