Mini-chromosome maintenance (MCM) proteins form a conserved family found in all eukaryotes and are essential for DNA replication. They exist as heteromultimeric complexes containing as many as six different proteins. These complexes are believed to be the replicative helicases, functioning as hexameric rings at replication forks. In most archaea a single MCM protein exists. The protein from Methanobacterium thermoautotrophicum (mtMCM) has been reported to assemble into a large complex consistent with a dodecamer. We show that mtMCM can assemble into a heptameric ring. This ring contains a C-terminal helicase domain that can be fit with crystal structures of ring helicases and an N-terminal domain of unknown function. While the structure of the ring is very similar to that of hexameric replicative helicases such as bacteriophage T7 gp4, our results show that such ring structures may not be constrained to have only six subunits.

The unique cytokine interleukin-18 (IL-18) acts synergistically with IL-12 to regulate T-helper 1 and 2 lymphocytes and, as such, seems to underlie the pathogenesis of various autoimmune and allergic diseases. Several anti-IL-18 agents are in clinical development, including the recombinant human antibody ABT-325, which is entering trials for autoimmune diseases. Given competing cytokine/receptor and cytokine/receptor decoy interactions, understanding the structural basis for recognition is critical for effective development of anti-cytokine therapies. Here we report three crystal structures: the murine antibody 125-2H Fab fragment bound to human IL-18, at 1.5 Å resolution; the 125-2H Fab (2.3 Å ); and the ABT-325 Fab (1.5 Å ). These structures, along with human/mouse IL-18 chimera binding data, allow us to make three key observations relevant to the biology and antigenic recognition of IL-18 and related cytokines. First, several IL-18 residues shift dramatically (>10 Å ) upon binding 125-2H, compared with unbound IL-18 (Kato, Z., Jee, J., Shikano, H., Mishima, M., Ohki, I., Ohnishi, H., Li, A., Hashimoto, K., Matsukuma, E., Omoya, K., Yamamoto, Y., Yoneda, T., Hara, T., Kondo, N., and Shirakawa, M. (2003) Nat. Struct. Biol. 10, 966 -971). IL-18 thus exhibits plasticity that may be common to its interactions with other receptors. Related cytokines may exhibit similar plasticity. Second, ABT-325 and 125-2H differ significantly in combining site character and architecture, thus explaining their ability to bind IL-18 simultaneously at distinct epitopes. These data allow us to define the likely ABT-325 epitope and thereby explain the distinct neutralizing mechanisms of both antibodies. Third, given the high 125-2H potency, 10 well ordered water molecules are trapped upon complex formation in a cavity between two IL-18 loops and all six 125-2H complementarity-determining regions. Thus, counterintuitively, tight and specific antibody binding may in some cases be water-mediated. Interleukin (IL)3 -18 is a proinflammatory cytokine that participates in the regulation of innate and acquired immunity (2, 3). IL-18 acts alone or in concert with IL-12 to amplify the induction of proinflammatory and cytotoxic mediators, such as interferon-␥. For example, in IL-18 knock-out mice, levels of interferon-␥ and cytotoxic T cells decrease despite the presence of IL-12. Inhibition of IL-18 activity has been found to be beneficial in several autoimmune disease animal models (e.g. collagen-induced arthritis (4) and colitis (5)). Furthermore, IL-18 expression is dramatically increased by the chronic inflammatory state extant in human autoimmune diseases, such as rheumatoid arthritis (6), multiple sclerosis (7, 8), and Crohn's disease (9). These observations suggest that blockade of IL-18 may be a useful human therapeutic modality (10).Despite functional divergence from the IL-1 cytokine family, IL-18 shares many similarities with IL-1. First, human IL-18 is synthesized as a biologically inactive 24-kDa precursor. Like IL-1␤, IL-18 is act...

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