SUNY at Albany
June 19-23, 2001
Archaeal histone-fold protein HMf creates bona fide chromatin fibers
Archaea, the third domain of life recently recognized (Woese et al., 1990), comprises microorganisms that are morphologically similar to prokaryotes, but possess many molecular features that put them closer to eukaryotes. Thus, understanding the structural and functional organization of their genomes seems to be an important clue to how these organisms handle their genetic information, and to how they may have evolved.
Archaea contain a family of small basic proteins that are structural homologues of eukaryal core histones H3 and H4. They contain the histone fold and dimerize in solution using the ÔhandshakeÕ interaction motif (Arents et al., 1991). Earlier reports have suggested that these proteins (HMf in particular) may organize DNA in the form of chromatin (reviewed in Reeve et al., 1997; Zlatanova, 1997).
Here, we have used the Atomic Force Microscope to image nucleosomal arrays reconstituted on the 208 bp tandemly repeated sequence from the 5S rDNA (Simpson et al., 1985) and three different proteins/protein complexes: the eukaryal histone octamer, the H3/H4 tetramer, with HMf from Methanothermus fervidis. We have also imaged nucleoprotein complexes formed on mononucleosome-sized DNA fragments. The AFM images were used to measure center-to-center internucleosomal distances, heights of individual particles, and the number of bp hidden within each type of particle. The AFM data was complemented by results from more conventional biochemical analyses, like methidiumpropyl-EDTA-iron(II) cleavage, micrococcal and restriction nuclease digestion, and band-shift analysis. We conclude that although HMf forms mononucleosomal particles that are much less stable than the canonical eukaryal nucleosomes, their internal structure is similar to that of H3/H4-containing nucleosomes. The structure of the HMf-fiber is also very similar to that of the eukaryal nucleosomal arrays, again strongly resembling that of the H3/H4 containing fibers. Thus, HMf is capable of organizing long stretches of DNA in the form of bona fide chromatin fibers.References and Footnotes
Tomschik, M., Karymov, M.A., Zlatanova, J. and Leuba, S.H.
Physical Molecular Biology (M.T., M.A.K.,S.H.L.),NCI, NIH, Bethesda, MD 20892
Chemistry and Chemical Engineering (J.Z.), Polytechnic U., Brooklyn, NY 11201