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Recent X-ray crystal structure determinations of Moloney murine leukemia virus reverse transcriptase (MoMLV RT) have allowed for more accurate structure/function comparisons to HIV-1 RT than were formerly possible. Previous biochemical studies of MoMLV RT in conjunction with knowledge of sequence homologies to HIV-1 RT and overall fold similarities to RTs in general, provided a foundation upon which to build. In addition, numerous crystal structures of the MoMLV RT fingers/palm subdomain had also shed light on one of the critical functions of the enzyme, specifically polymerization. Now in the advent of new structural information, more intricate examination of MoMLV RT in its entirety can be realized, and thus the comparisons with HIV-1 RT may be more critically elucidated. Here, we will review the similarities and differences between MoMLV RT and HIV-1 RT via structural analysis, and propose working models for the MoMLV RT based upon that information.

The properties of high-pH suspensions of mixtures of silica with low-molecular-weight samples of the water-soluble polymer polyethylenimine (PEI) have been studied. At pH > 10 and low ionic strength, silica nanoparticles are stabilized by a negative surface charge, and PEI has only a very low positive charge. The adsorption of PEI induces a localized positive charge on the segments of polymer closest to the silica surface. The parts of the molecule furthest away from the surface have little charge because of the high pH of the medium. The polymer-covered particle remains negatively charged, imparting some electrostatic stabilization. Suspensions of silica and low-molecular-weight PEI are low-viscosity fluids immediately after mixing, but aggregation occurs leading to the eventual gelation (or sedimentation at lower concentrations) of these mixtures, indicating colloidal instability. The gelation time passes through a minimum with increasing surface coverage. The rate of gelation increases exponentially with molecular weight: for molecular weight> or = 10,000 Da PEI, the instability is so severe that uniform suspensions cannot be produced using simple mixing techniques. The gelation rates increase rapidly with temperature, ionic strength, and reduction in pH. The rate of gelation increases with increasing particle concentration at low surface coverage but decreases at high coverage as a consequence of a small increase in pH. Gels are broken by application of high shear into aggregates that re-gel more rapidly than the original discrete coated particles.

Ndt80 is a transcriptional activator required for meiosis in the yeastSaccharomyces cerevisiae. Here, we report the crystal structure at 2.3 Å resolution of the DNA-binding domain of Ndt80 experimentally phased by using the anomalous and isomorphous signal from a single ordered Se atom per molecule of 272-aa residues. The structure reveals a single Ϸ32-kDa domain with a distinct fold comprising a ␤-sandwich core elaborated with seven additional ␤-sheets and three short ␣-helices. Inspired by the structure, we have performed a mutational analysis and defined a DNA-binding motif in this domain. The DNA-binding domain of Ndt80 is homologous to a number of proteins from higher eukaryotes, and the residues that we have shown are required for DNA binding by Ndt80 are highly conserved among this group of proteins. These results suggest that Ndt80 is the defining member of a previously uncharacterized family of transcription factors, including the human protein (C11orf9), which has been shown to be highly expressed in invasive or metastatic tumor cells.transcription factor ͉ sporulation ͉ meiotic ͉ ␤-sandwich

Complexation with the N-terminal fragment of Moloney murine leukemia virus reverse transcriptase offers a novel method of obtaining crystal structures of nucleic acid duplexes, which can be phased by molecular replacement. This method is somewhat similar to the method of using a monoclonal antibody Fab fragment complexed to the molecule of interest in order to obtain crystals suitable for X-ray crystallographic analysis. Here a novel DNA structure including two G±A mispairs in a pseudo-hexadecamer determined at 2.3 A Ê resolution in a complex with the N-terminal fragment is reported. This structure has an asymmetric unit consisting of the protein molecule bound to the blunt end of a DNA 6/10-mer, which is composed of a six-base strand (5 H -CTCGTG-3 H ) and a ten-base strand (3 H -GAGCACGGCA-5 H ). The 6/10-mer is thus composed of a six-base-pair duplex with a four-base single-stranded overhang. In the crystal structure, the bases of the overhang are reciprocally paired (symmetry element Àx À 1, Ày, z), yielding a doubly nicked pseudo-hexadecamer primarily B-form DNA molecule, which has some interesting A-like structural features. The pairing between the single strands results in two standard (G±C) Watson±Crick pairs and two G±A mispairs. The structural DNA model can accommodate either a standard syn or a standard anti conformation for the 5 H -terminal adenine of the ten-base strand of the DNA based on analysis of simulated-annealing omit maps. Although the DNA model here includes nicks in the phosphodiester backbone, modeling of an intact phosphodiester backbone results in a very similar DNA model and indicates that the structure is biologically relevant.