Distamycin and Hoechst 33258 have long served as the model compounds for biochemical, biophysical, and clinical studies of the drugs that bind in the DNA minor groove. However, the results presented in this investigation clearly show that 4,6-diamidino-2 phenylindole (DAPI) is superior to both of these drugs at negating the effects of intrinsic DNA curvature and anisotropic bendability as measured by electrophoretic and ligation analysis. In addition, DAPI was more effective than distamycin and Hoechst 33258 at inhibiting the assembly of nucleosomes onto synthetic and natural sequences that have multiple closely spaced oligo-AT sequences that serve as drug binding sites. Since these effects may be related to the biological action of the drugs, it was of interest to determine the mechanism that was responsible for the enhanced action of DAPI. The possibility that the differential drug potencies resulted from differential overall affinities of the ligands for A-tract molecules was considered, but drug binding studies suggested that this was not the case. It is also unlikely that the differential drug effects resulted from the binding of the drugs to different DNA sites since the oligo A/T binding sites for DAPI and Hoechst were centered on the same nucleotide positions as revealed by footprinting studies using exonuclease III, DNase I, and hydroxyl radical. However, the footprinting studies with DNase I did uncover a potentially important difference between the drugs. DAPI protected only the AT bp in the binding sites, while distamycin and Hoechst protected these bp as well as flanking Gs and Cs. These results permitted us to advance a preliminary model for the enhanced action DAPI. According to the model, the short length of DAPI and its absolute specificity for A/T bps with narrow minor grooves ensures that only particularly minor grooves that give rise to curvature and anisotropic bendability are occupied by the drug. Consequently, each helical deflection induced by an A-tract in the absence of the drug is countered by an opposite deflection induced by DAPI binding, thus effectively neutralizing intrinsic curvature and bending into the minor groove.

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Abstract. Rock and soil samples from the planet Mars are due to be returned to Earth within a decade. Martian samples initially will be tested for evidence of life and biological hazard under strict biological containment. Wider distribution of samples for organic and inorganic analysis may occur only if neither evidence of life nor hazard is detected, or if the samples are first sterilized. We subjected a range of Mars analog rocks and minerals to high doses of gamma radiation in order to determine the effects of gamma sterilization on the samples' isotopic, chemical, and physical properties. Gamma photons from 6øCo (1.17 and 1.33 MeV) in doses as high as 3 x 107 rads did not induce radioactivity in the samples and produced no measurable changes in their isotopic and chemical compositions. This level of irradiation also produced no measurable changes in the crystallographic structure of any sample, the surface areas of soil analogs, or the fluid inclusion homogenization temperature of quartz. The only detectable effects of irradiation were dose-dependent changes in the visible and near-infrared spectral region (e.g., discoloration and darkening of quartz and halite and an increase in albedo of carbonates) and increases in the thermoluminescence of quartz and plagioclase. If samples returned from Mars require biological sterilization, gamma irradiation provides a feasible option. BackgroundMartian rock and soil, collected by robotic spacecraft, will be returned to terrestrial laboratories early in the next century. The return of documented samples, carefully collected and preserved, will be a major step in the search for evidence of Martian life. Martian conditions, including the lack of organic matehal, subfreezing temperatures, high flux of solar ultraviolet radiation, and strongly oxidizing chemical species in the soil, severely limit the survival of organisms at or near the surface [Klein, 1998] inCenters for Disease Control and Prevention, Atlanta, Georgia.•University of Portsmouth, Portsmouth, England.Copyright 1999 by the American Geophysical Union. Paper number 1999JE001064.0148-0227/99/1999JE001064509.00 specialized organisms at low concentrations in the returned samples is conceivable [Clark, 1998].Current planetary protection strategies call for the samples to be immediately placed into biological containment and tested for signs of present or past life and biological hazard [DeVincenzi et al., 1998]. It is recommended that "Controlled distribution of unsterilized materials from Mars should occur only if rigorous analyses determine that the materials do not constitute a biological hazard. If any portion of the sample is removed from containment prior to completion of these analyses it should first be sterilized" [Space Studies Board, 1997]. While sterilization of Mars samples may not be required, an acceptable method must be available before the samples are returned to Earth.A variety of sterilization techniques have been used or proposed for spacecraft missions to Mars. These include dry heating to t...

A computer program for predicting DNA bending from nucleotide sequence was used to identify circular structures in retroviral and cellular genomes. An 830-base pair circular structure was located in a control region near the center of the genome of the human immunodeficiency virus type I (HIV-I). This unusual structure displayed relatively smooth planar bending throughout its length. The structure is conserved in diverse isolates of HIV-I, HIV-II, and simian immunodeficiency viruses, which implies that it is under selective constraints. A search of all sequences in the GenBank data base was carried out in order to identify similar circular structures in cellular DNA. The results revealed that the structures are associated with a wide range of sequences that undergo recombination, including most known examples of DNA inversion and subtelomeric translocation systems. Circular structures were also associated with replication and transposition systems where DNA looping has been implicated in the generation of large protein-DNA complexes. Experimental evidence for the structures was provided by studies which demonstrated that two sequences detected as circular by computer preferentially formed covalently closed circles during ligation reactions in vitro when compared to nonbent fragments, bent fragments with noncircular shapes, and total genomic DNA. In addition, a single T 3 C substitution in one of these sequences rendered it less planar as seen by computer analysis and significantly reduced its rate of ligase-catalyzed cyclization. These results permit us to speculate that intrinsically circular structures facilitate DNA looping during formation of the large protein-DNA complexes that are involved in site-and region-specific recombination and in other genomic processes.Sequence-directed bending of DNA is one factor that causes local variations in the structure of genomes (reviewed by

An understanding of virus disassembly requires a detailed understanding of the protein-protein and protein-nucleic acid interactions which stabilize the virion. We have characterized a mutant of cowpea chlorotic mottle virus [cpR26C (coat protein R26C)] that displays increased virion stability and is abnormal in virion disassembly when purified under nonreducing conditions. Reduced virions are infectious, whereas nonreduced virions are noninfectious. The cpR26C mutant virions purified under nonreducing conditions resist disassembly in 0.5 M CaCl2, pH 7.5. The nonreduced cpR26C mutant virions swell in neutral pH conditions (pH 7.5) but do not disassociate when the ionic strength is increased. In contrast, wild-type virions or cpR26C mutant virions isolated under reducing conditions completely disassociate into the RNA and capsid protein components at pH 7.5 and high ionic strength (i > 1.0). Sequence analysis of the cpR26C mutant identified a single C to U nucleotide change at position 1435 of RNA 3 (position 86 of RNA 4), which results in a arginine to cysteine change at position 26 of the coat protein. The cpR26C mutant provides an ideal chemical switch for examining virion assembly and disassembly.