Most patients with juvenile periodontitis manifest serum antibodies, sometimes at very high titers, to antigens of ActinobaciUus actinomycetemcomitans, but the antigens inducing the immune response have been only partly characterized. We separated A. actinomycetemcomitans serotype b cells into protein, lipopolysaccharide (LPS), and soluble polysaccharide fractions and characterized them. Coomassie blueand silver-stained sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels were used to detect protein and LPS components, and gas-liquid chromatography was used to determine their carbohydrate and fatty acid composition. Western blots, dot blots, and enzyme-linked immunosorbent assay inhibition with high-titer sera from juvenile periodontitis patients revealed which components were highest in antibody binding activity. These results showed that the major portion of the immunoglobulin G binding activity resides in the purified mannan-free LPS, with lesser amounts in the total protein fraction. Using Sephacryl S-300 chromatography, we separated LPS into high-molecular-mass components with high carbohydrate contents by gas-liquid chromatography and a low-molecular-mass component consisting mainly of lipid A and the inner core sugar heptulose. The results of quantitative dot blot assays and enzyme-linked immunosorbent assay inhibition show that the serotype-specific antibody binding activity is highly concentrated in the high-molecular-mass carbohydrate-rich LPS fraction and is almost completely absent in the low-molecular-weight lipid-rich fraction. Our observations contrast with previous reports that the predominant serotype antigen of A. actinomycetemcomitans resides in a mannan-rich polysaccharide isolated from spent culture medium. These observations support the conclusion that the immunodominant antigen of the outer membrane is the 0 antigen of the LPS.

Smc2/4 forms the core of the Saccharomyces cerevisiae condensin, which promotes metaphase chromosome compaction. To understand how condensin manipulates DNA, we used two in vitro assays to study the role of SMC (structural maintenance of chromosome) proteins and ATP in reconfiguring the path of DNA. The first assay evaluated the topology of knots formed in the presence of topoisomerase II. Unexpectedly, both wild-type Smc2/4 and an ATPase mutant promoted (؉) chiral knotting of nicked plasmids, revealing that ATP hydrolysis and the non-SMC condensins are not required to compact DNA chirally. The second assay measured Smc2/4-dependent changes in linking number (Lk). Smc2/4 did not induce (؉) supercoiling, but instead induced broadening of topoisomer distributions in a cooperative manner without altering Lk 0 . To explain chiral knotting in substrates devoid of chiral supercoiling, we propose that Smc2/4 directs chiral DNA compaction by constraining the duplex to retrace its own path. In this highly cooperative process, both (؉) and (؊) loops are sequestered (about one per kb), leaving net writhe and twist unchanged while broadening Lk. We have developed a quantitative theory to account for these results. Additionally, we have shown at higher molar stoichiometries that Smc2/4 prevents relaxation by topoisomerase I and nick closure by DNA ligase, indicating that Smc2/4 can saturate DNA. By electron microscopy of Smc2/4-DNA complexes, we observed primarily two protein-laden bound species: long flexible filaments and uniform rings or "doughnuts." Close packing of Smc2/4 on DNA explains the substrate protection we observed. Our results support the hypothesis that SMC proteins bind multiple DNA duplexes.SMC (structural maintenance of chromosome) proteins are the central components of several multiprotein complexes that help to organize chromosomes throughout the cell cycle (1-5). Understanding the properties of the SMC proteins may illuminate their essential function in cohesin, condensin, "compensin," repair complexes containing Smc5/6, and recombination complexes (6 -10). Disruption of SMC function in prokaryotes and eukaryotes alike leads to chromosome instability, defects in repair and recombination, and death under conditions of rapid growth (11-15). The budding yeast Smc2 and Smc4 proteins form a heterodimeric ATPase (16) that, in combination with the non-SMC proteins Brn1, Ycg1, and Ycs4, comprises the holo-condensin enzyme. Holo-condensin drives global chromosome condensation at mitosis (4, 17, 18) and specific condensation of the rDNA locus at anaphase (19). The Xenopus 8 S condensin FF and the Schizosaccharomyces pombe Cut3/14 complex are examples of other stable condensin SMC pairs whose properties have been studied in some detail (20 -23).The SMC proteins are large (ϳ150 kDa) and contain five structural motifs that include N-and C-terminal globular domains, a central hinge domain (H), and two long ␣-helices (␣ N and ␣ C ) arranged as follows: N-␣ N -H-␣ C -C. Coiled-coil pairing of ␣ N and ␣ C folds a single...

As genetic material DNA is wonderful, but as a macromolecule it is unruly, voluminous and fragile. Without the action of DNA replicases, topoisomerases, helicases, translocases and recombinases, the genome would collapse into a topologically entangled random coil that would be useless to the cell. We discuss the organization, movement and energetics of these proteins that are crucial to the preservation of a molecule that has such beautiful biological but challenging physical properties.

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