An unusual presentation of a linear epidermal nevus.

We report an unusual presentation of a linear epidermal nevus in an adolescent male. Epidermal nevi most commonly appear in infancy and early childhood. They often are found in association with other organ system anomalies. We describe a palmar linear epidermal nevus that caused impairment of the patient's use of his hand. Therapeutic management involved surgical excision of the nevus and reconstruction of the area with a full-thickness skin graft.

Role of macromolecular hydration in the binding of the Escherichia coli cyclic AMP receptor to DNA.

The osmotic stress technique was used to measure the changes in macromolecular hydration that accompany binding of the Escherichia coli CAP protein to its transcription-regulatory site (C1) in the lactose promoter and that accompany the transfer of CAP from site C1 to nonspecific genomic DNA. Formation of the C1 complex is accompanied by the net release of 79 +/- 11 water molecules. If all water molecules were released from macromolecular surfaces, this result would be consistent with a net reduction of solvent-accessible surface area of 711 +/- 189 A2. This area is only slightly smaller than the solvent-inaccessible macromolecular interface in crystalline CAP-DNA complexes. The transfer of CAP from site C1 to nonspecific sites is accompanied by the net uptake of 56 +/- 10 water molecules. Taken with the water stoichiometry of sequence-specific binding, this value implies that formation of a nonspecific complex is accompanied by the net release of 2-44 water molecules. The enhanced stabilities of CAP-DNA complexes with increased osmolality (decreased water activity) may contribute to the ability of E.coli cells to tolerate dehydration and/or high external salt concentrations.

Sequestration stabilizes lac repressor-DNA complexes during gel electrophoresis.

The gel electrophoresis mobility shift assay is widely used for both qualitative and quantitative characterization of protein-nucleic acid interactions. Often it is found that protein-nucleic acid complexes persist within gels for much longer than would be expected on the basis of their free solution lifetimes. Excluded volume and matrix-interaction mechanisms have been proposed to account for the enhanced stabilities of complexes within gels. To test these mechanisms, we have investigated the influences of gel composition and concentration on the pseudo first-order dissociation kinetics of complexes containing the Escherichia coli lactose (lac) repressor protein and lactose promoter DNA. In both polyacrylamide and agarose gels, dissociation rates were slower than those in free solution and decreased with increasing gel concentration. This result is inconsistent with mechanisms of stabilization that require specific interactions with the gel matrix. Under standard reaction conditions, free solution values of kdiss were proportional to [DNA]0.83 +/- 0.11, while in 10% polyacrylamide gels kdiss values were proportional to [DNA]0.48 +/- 0.09. These results suggest that the lifetimes of lac repressor-DNA complexes in free solution are limited by their encounter frequency with molecules of DNA or with protein-DNA complexes; some or all of the stabilization observed in gels may be due to a reduction in this frequency.

The mechanism of CAP-lac repressor binding cooperativity at the E. coli lactose promoter.

The cyclic AMP receptor protein (CAP) and lactose repressor bind their regulatory sites in the lactose promoter with moderate cooperativity (omega C101 = 11.8(+/- 3.7)). This cooperativity is significantly reduced by the removal of DNA located upstream of the CAP binding site or by substitution of the dimeric lacI-18 mutant repressor for the wild-type tetrameric protein. These results are consistent with a mechanism of interaction in which CAP bends the DNA and the lac repressor binds simultaneously to its operator site and to promoter-distal sequences. Similar values of omega C101 were obtained with a promoter truncation containing the O3 pseudooperator site and one in which the site is destroyed, suggesting that DNA contacts distal to the O3 site are necessary for cooperative binding.

Free DNA concentration in E. coli estimated by an analysis of competition for DNA binding proteins.

Transcription in E. coli is often controlled by the binding of specific gene-regulatory proteins. Binding of these proteins to their specific DNA binding sites occurs in the presence of a large excess of "nonspecific" genomic DNA. Binding to a specific DNA site thus depends on the concentration of regulatory protein, on its affinities for specific and competing nonspecific binding sites, and on the free concentrations of those sites. Although it is probable that genomic DNA is largely occluded by protein binding or by condensation in vivo, the actual extent to which the DNA is available to act as a competitor for specific binding (i.e. the effective concentration of nonspecific DNA) is not known. Because many regulatory interactions occur simultaneously in a cell, it is reasonable to expect that they will have evolved to function at equilibrium with a shared concentration of competing nonspecific DNA. This premise was the basis for this study. In vitro binding data were compiled for six regulatory proteins that function in E. coli, and used to calculate theoretical equilibrium binding distributions. The calculated distributions were used to evaluate the regulatory states of promoters according to models based on the equilibrium occupancies of regulatory sites. For four proteins whose DNA-binding affinities are modulated by ligand binding (CAP, lac repressor, trp repressor and araC), regulation was assessed as the extent to which the presence of the modulator could affect the occupancy by protein of the specific sites (e.g. the difference in equilibrium occupancy by CAP of CAP binding sites between conditions of high and low concentrations of CAP's affinity modulator, cAMP). For two proteins whose site affinities are not modulated by ligand binding (lambda repressor and lambda-cro), regulation was assessed by specific site occupancy at equilibrium. These regulation profiles were compared to determine whether a single concentration of nonspecific competing DNA is compatible with effective regulation as defined for all of the systems. For five of the six modeled systems (CAP, trp repressor, araC, lambda repressor and lambda-cro), a free nonspecific DNA concentration on the order of 10(-4) M base pairs is compatible with regulation based on equilibria of the protein-DNA interactions. The lac repressor-operator system is an exception to these results: as has been shown previously, the regulation of operator binding by low molecular weight inducers increases with increasing concentrations of nonspecific DNA (von Hippel et al., 1974 Proc. natn. Acad. Sci. U.S.A. 71, 4808-4812).(ABSTRACT TRUNCATED AT 400 WORDS)