Eukaryotic chromosomal DNA is licensed for replication precisely once in each cell cycle. The mini-chromosome maintenance (MCM) complex plays a role in this replication licensing. We have determined the structure of a fragment of MCM from Methanobacterium thermoautotrophicum (mtMCM), a model system for eukaryotic MCM. The structure reveals a novel dodecameric architecture with a remarkably long central channel. The channel surface has an unusually high positive charge and binds DNA. We also show that the structure of the N-terminal fragment is conserved for all MCMs proteins despite highly divergent sequences, suggesting a common architecture for a similar task: gripping/remodeling DNA and regulating MCM activity. An mtMCM mutant protein equivalent to a yeast MCM5 (CDC46) protein with the bob1 mutation at its N terminus has only subtle structural changes, suggesting a Cdc7-bypass mechanism by Bob1 in yeast. Yeast bypass experiments using MCM5 mutant proteins support the hypothesis for the bypass mechanism.

High risk human papillomavirus types 16 (HPV16) and 18 (HPV18) can cause cervical cancer. Efficient infection by HPV16 and HPV18 pseudovirions requires interactions of particles with cell-surface receptor heparan sulfate oligosaccharide. To understand the virus-receptor interactions for HPV infection, we determined the crystal structures of HPV16 and HPV18 capsids bound to the oligosaccharide receptor fragment using oligomeric heparin. The HPV-heparin structures revealed multiple binding sites for the highly negatively charged oligosaccharide fragment on the capsid surface, which is different from previously reported virus-receptor interactions in which a single type of binding pocket is present for a particular receptor. We performed structure-guided mutagenesis to generate mutant viruses, and cell binding and infectivity assays demonstrated the functional role of viral residues involved in heparin binding. These results provide a basis for understanding virus-heparan sulfate receptor interactions critical for HPV infection and for the potential development of inhibitors against HPV infection. Human papillomaviruses (HPVs)4 are non-enveloped small DNA viruses of great medical importance. Among the large group of HPVs known by now, sexually transmitted genital high risk HPV types are the cause for the development of a variety of epithelial tumors, especially cervical carcinoma (1). Cervical cancer is the second leading cause of death among female cancer patients worldwide. HPV16 and HPV18 stand out, as they are causally linked to Ͼ70% of cervical cancer cases (2).HPV particles consist of 72 pentamers of the major capsid protein L1, which forms the virus outer shell and encapsidates the viral DNA (3, 4). The minor capsid protein L2 is present at up to 72 copies and is hidden inside the capsid with exception of a small N-terminal section (5, 6). Efficient infection by HPV16 and HPV18 pseudoviruses requires the interactions of the L1 protein with extracellular matrix (ECM)-and cell surface-resident heparan sulfate receptor in vitro (7-9) as well as in vivo models (10). Homologs of heparan sulfate polysaccharide or heparin, secreted by mast cells, can inhibit HPV infection (7-9).Cell-surface heparan sulfates are linear and highly negatively charged oligosaccharides that are covalently linked to proteins. They can serve as the attachment receptors for several important human virus pathogens (7,11,12). Despite considerable efforts, the interactions between HPV and the heparan sulfate oligosaccharides that initiate infection are poorly understood. Here, we determined the co-crystal structure of HPV16 and HPV18 capsids bound to oligomeric heparin. We found that the highly negatively charged heparin fragment binds to multiple locations on the capsid surface mainly through charge-charge interactions. On the basis of the structure, we generated mutant virus to disrupt the interactions with heparin. ECM and cell binding assays combined with infectivity measurements showed that substitution of key HPV residues involved in bi...

Human papillomaviruses (HPVs) are known etiologic agents of cervical cancer. Vaccines that contain virus-like particles (VLPs) made of L1 capsid protein from several high risk HPV types have proven to be effective against HPV infections. Raising high levels of neutralizing antibodies against each HPV type is believed to be the primary mechanism of protection, gained by vaccination. Antibodies elicited by a particular HPV type are highly specific to that particular HPV type and show little or no cross-reactivity between HPV types. With an intention to understand the interplay between the L1 structure of different HPV types and the type specificity of neutralizing antibodies, we have prepared the L1 pentamers of four different HPV types, HPV11, HPV16, HPV18, and HPV35. The pentamers only bind the type-specific neutralizing monoclonal antibodies (NmAbs) that are raised against the VLP of the corresponding HPV type, implying that the surface loop structures of the pentamers from each type are distinctive and functionally active as VLPs in terms of antibody binding. We have determined the crystal structures of all four L1 pentamers, and their comparisons revealed characteristic conformational differences of the surface loops that contain the known epitopes for the NmAbs. On the basis of these distinct surface loop structures, we have provided a molecular explanation for the type specificity of NmAbs against HPV infection.