In contrast to the cell division machineries of bacteria, euryarchaea, and eukaryotes, no division components have been identified in the second main archaeal phylum, Crenarchaeota. Here, we demonstrate that a three-gene operon, cdv, in the crenarchaeon Sulfolobus acidocaldarius, forms part of a unique cell division machinery. The operon is induced at the onset of genome segregation and division, and the Cdv proteins then polymerize between segregating nucleoids and persist throughout cell division, forming a successively smaller structure during constriction. The cdv operon is dramatically down-regulated after UV irradiation, indicating division inhibition in response to DNA damage, reminiscent of eukaryotic checkpoint systems. The cdv genes exhibit a complementary phylogenetic range relative to FtsZ-based archaeal division systems such that, in most archaeal lineages, either one or the other system is present. Two of the Cdv proteins, CdvB and CdvC, display homology to components of the eukaryotic ESCRT-III sorting complex involved in budding of luminal vesicles and HIV-1 virion release, suggesting mechanistic similarities and a common evolutionary origin.he Archaea constitute a separate domain of life that has evolved in parallel with Bacteria and Eukarya (1). The archaeal domain is currently divided into two main lineages, the Crenarchaeota and the Euryarchaeota, each of which comprises several distinct classes of organisms that thrive in a wide variety of environments. Whereas several aspects of archaeal biology appear to be unique, certain traits resemble those in eukaryotes, including the machineries that govern information storage, maintenance, and processing.Several features of archaeal cell cycle progression have been elucidated in considerable detail including the overall organization of the cell cycle in certain species, and regulatory and mechanistic aspects of the replication process (2, 3). Conversely, the genome segregation machinery remains essentially uncharacterized in this domain. In archaeal species belonging to the Euryarchaeota phylum, and in bacteria, cell division is mediated by FtsZ protein filaments that form a constricting ring structure (4). In eukaryotes, division occurs with the help of a contractile actin-myosin ring or, in plant cells, by septum formation at a site initially marked by actin and microtubules (5). In contrast to bacteria, euryarchaea, and eukaryotes, no cell division components have been identified in the second main archaeal phylum, Crenarchaeota (2).Here, we report on the identification of key components of the cell division system in the hyperthermophilic crenarchaeon Sulfolobus acidocaldarius, describe intracellular structures that are formed by the gene products during genome segregation and division, and show that the operon is subject to a checkpoint-like regulation. We also demonstrate that the division machinery is present in all crenarchaeal orders except Thermoproteales, and that it is related to the eukaryotic ESCRT-III sorting complex.

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The interactions of a set of compounds of potential importance for anticancer and AIDS chemotherapy with lipid membranes and plasma proteins were studied with a surface plasmon resonance (SPR) based optical biosensor, giving valuable information on the absorption and distribution of the compounds. The technique allowed both effective screening of compounds and more detailed kinetic and mechanistic analysis of specific interactions. The interaction with two different types of lipid membranes could be reliably measured at a drug concentration as low as 20 microM, allowing analysis of poorly soluble compounds. Distribution was evaluated by investigation of the interactions with two human plasma proteins, human serum albumin (HSA) and alpha(1)-acid glycoprotein (AGP). Two apparent binding sites were clearly defined for HSA: one with rapid and one with slow association and dissociation rates. The sites appear to differ in accessibility and recognition characteristics rather than in their capacities to form strong complexes with drugs.

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Surface plasmon resonance biosensor technology (SPR) is ideally suited for fragment-based lead discovery. However, generally suitable experimental procedures or detailed protocols are lacking, especially for structurally or physico-chemically challenging targets or when tool compounds are lacking. Success depends on accounting for the features of both the target and the chemical library, purposely designing screening experiments for identification and validation of hits with desired specificity and mode-of-action, and availability of orthogonal methods capable of confirming fragment hits. By adopting a multiplexed strategy, the range of targets and libraries amenable to an SPR biosensor-based approach for identifying hits is considerably expanded. We here illustrate innovative strategies using five challenging targets and variants thereof. Two libraries of 90 and 1056 fragments were screened using two different flow-based SPR biosensor systems, allowing different experimental approaches. Practical considerations and procedures accounting for the characteristics of the proteins and libraries, and that increase robustness, sensitivity, throughput and versatility are highlighted.