crown@dukempd.UUCP (Rick Crownover) (08/25/88)
In reading "The Cell" by Alberts et. al. I ran across an 'open question' about chromatin packing (ch. 8 pp 389). I have a naive experimental approach to this question for discussion... but don't want to flog a dead horse. The book is a couple of years old now; so, does anyone know whether this question has been resolved in the meantime? Thanks, Rick -- Rick Crownover 1-919-684-8279 Duke University Dept. of Physics crown@dukempd.uucp Durham, N.C. 27706 mcnc!duke!dukempd!crown
eddy@boulder.Colorado.EDU (Sean Eddy) (08/28/88)
In article <707@dukempd.UUCP> crown@dukempd.UUCP (Rick Crownover) writes: > In reading "The Cell" by Alberts et. al. I ran across an >'open question' about chromatin packing (ch. 8 pp 389). I have >a naive experimental approach to this question for discussion... >but don't want to flog a dead horse. The book is a couple of years >old now; so, does anyone know whether this question has been >resolved in the meantime? Thanks, Rick Resolved, in favor of the drawing to the right (nucleosome hockey-pucks stacked on end rather than on their sides). Experimental technique involved X-ray diffraction of chromatin fibers. The two models for packing predicted different and distinguishable sets of reflections (which I don't understand anyway). I can look up a reference if you want. What was your experiment?? - Sean Eddy - Molecular/Cellular/Developmental Biology; U. of Colorado at Boulder - eddy@boulder.colorado.EDU !{hao,nbires}!boulder!eddy - - "Truth emerges more readily from error than from confusion." - - Francis Bacon
crown@dukempd.UUCP (Rick Crownover) (08/29/88)
The experiment I had in mind would have required synthesizing
a chain of known (and hopefully repetitive) sequence. This chain
would then be placed in solution with the histones and allowed to
pack up (Albert's text suggested that this would occur spontaneously).
After packing, a small molecule with ends capable of bonding to
specific AA's (which have been included in the original chain at
known intervals) would be introduced in a quantity large enough to
saturate the appropriate sites on the chain. Picture this second
molecule as forming "bridges" between two points when possible, or
attaching like a branch on a tree when it can't reach a second
suitable site.
Now denature the packed chromatin and cleave the chain with a
bond specific 'something-ase' which will break the chain -- again
in known positions along the chain. At this point a determination
of molecular weights for the cleavage residues (terminology???)
would be performed. By comparing the experimental weights with a
statistical model based expectations for the two possible structures,
it should be possible to select one or the other.
If the chemistry is up to this, the statistical model is easy.
X-ray crystallography is probably simpler though... and if its been
done... Anyway, that was the idea. Next.
--
Rick Crownover 1-919-684-8279
Duke University Dept. of Physics crown@dukempd.uucp
Durham, N.C. 27706 mcnc!duke!dukempd!crowneddy@boulder.Colorado.EDU (Sean Eddy) (08/29/88)
In article <709@dukempd.UUCP> crown@dukempd.UUCP (Rick Crownover) writes:
[design of a cross-linking study to probe chromatin structure]
This is somewhat similar to an approach I've heard rumor of, so
I'm going to take the opportunity to bring it up.
"DNA footprinting" is a popular technique for learning where a
protein binds to a particular piece of DNA. Basically, the
protein blocks modification of the DNA helix and protects
its binding site from chemical or enzymatic cleavage;
this "protection" can be detected and is called the "footprint"
of the protein.
Now, has anyone heard of _protein_ footprinting being used to probe
protein-protein or protein-nucleic acid interactions?
Would a protein-protein interaction
protect appropriate peptide bonds from cleavage by, say, a protease?
Would a protein make a protease-insensitive "footprint" on another
protein? (Like I said, I've heard rumor that the answers are yes;
anyone know anything about this?)
- Sean Eddy
- Molecular/Cellular/Developmental Biology; U. of Colorado at Boulder
- eddy@boulder.colorado.EDU !{hao,nbires}!boulder!eddy