Scott C. Mohr

An Estradiol-Porphyrin Conjugate Selectively Localizes Into Estrogen Receptor-Positive Breast Cancer Cells

A conjugate of a C(11)-beta-derivative of estradiol and an asymmetric tetraphenylporphyrin was synthesized to study its potential selective uptake by breast cancer cells naturally over-expressing the nuclear receptor for estrogen (ER). Competitive radioligand binding assays of this conjugate with recombinant ER showed that the conjugate bound to ER in a dose-dependent manner with an EC50 of 274 nM, compared with 1 nM for estradiol, the natural ligand. Cellular uptake studies with ER-positive MCF-7 and ER-negative HS578t human breast cancer cells revealed that, the conjugate was taken up by MCF-7 cells in a dose-dependent manner, which was obliterated by co-incubation with a large excess of estradiol. On the other hand there was very little uptake of the un-conjugated porphyrin by MCF-7 and Hs578t cells. HS578t cells also showed insignificant uptake of the conjugate under the conditions of our experiment. These results strongly suggested that specific interaction between the endogenous ER in MCF-7 cells and the estrogen part of the conjugate enabled these cells to selectively internalize the conjugate over the un-conjugated porphyrin. Therefore, ER-binding conjugates of estradiol and porphyrins could potentially be used for ER-targeted photodynamic therapy of hormone-sensitive cancers of breast, ovary, gonads etc.

The thermal transition of 'psi' DNA monitored by circular dichroism.

The Watson-Crick model for nucleic acid secondary structure fails to provide a basis for understanding the extreme degree of compactness exhibited by DNA molecules in vivo. Purified high-molecular-weight native DNA in vitro adopts a ‘stiff coil’ conformation whose dimensions even at high ionic strength far exceed those of the biological structure (virus particle, bacterial nucleoid, eukaryotic chromosome) from which the DNA was originally isolated [l-3]. The recent discovery by Lcrman [4] that above a critical ionic strength high concentrations of an inert macromolecule such as poly(ethylene oxide) (PEO) cause DNA to collapse into a rapidly sedimenting form strongly suggests that ‘exclusion interaction’ between DNA and other macromolecules plays a role in generating compact DNA conformations in vivo. Profound changes in the circular dichroism (CD) spectrum accompany this ‘psi’ or

What Rough Beast: Synthetic Biology, Uncertainty, and the Future of Biosecurity

(polymerand salt-induced) transition [5,6] and we have accordingly employed CD to investigate the thermal stability of the

Relaxation kinetics of the binding of ethidium bromide to unfractionated yeast tRNA at low dye/phosphate ratio☆

state of calf thymus DNA. Our results show that II/-DNA possesses a fairly sharp thermal transition whose mid-point (T,‘) which lies far below the helix-coil transition temperature (T,) depends strongly upon PEO concentration, but only weakly on ionic strength. Above the transition region the CD spectrum resembles that of the ordinary B-form DNA though with somewhat reduced intensity. This fact, combined with the reversibility of the transition, demonstrates contrary to an earlier report [7] that

Protein synthesis elongation factor 1 from rat liver: A zinc metalloenzyme

-DNA melts to give B-DNA, not denatured DNA. The melting behavior which we report here is

Transfer RNA: Change of conformation upon aminoacylation determined by Raman spectroscopy☆

Abstract Synthetic biology seeks to create modular biological parts that can be assembled into useful devices, allowing the modification of biological systems with greater reliability, at lower cost, with greater speed, and by a larger pool of people than has been the case with traditional genetic engineering. We assess the offensive and defensive security implications of synthetic biology based on the insights of leading synthetic biologists into how the technology may develop, the projections of practicing biosecurity authorities on changes in the security context and potential security applications of synthetic biology, and joint appraisals of policy relevant sources of uncertainty. Synthetic biology appears to have minimal security implications in the near term, create modest offensive advantages in the medium term, and strengthen defensive capabilities against natural and engineered biological threats and enable novel potential offensive uses in the long term. To maximize defensive and minimize offensive effects of synthetic biology despite uncertainty, this essay suggests a combination of policy approaches, including community-based efforts, regulation and surveillance, further research, and the deliberate design of security and safety features into the technology.

Survey of human mitochondrial diseases using new genomic/proteomic tools

Abstract Temperature-jump kinetic studies of the binding of ethidium bromide to unfractionated tRNA from yeast in the absence of Mg ++ reveal the presence of a three-step process. The first two steps are consistent with an intercalation mechanism as proposed by other workers to describe the interaction of planar polycyclic molecules with double helical nucleic acids. The third step appears to be unique for tRNA and we ascribe it to a change in the three dimensional conformation of the intercalated complex. Rate and equilibrium constants for the elementary steps have been calculated.

Why do we need a three-dimensional architecture of the ligand-binding domain of the nuclear 1α, 25-dihydroxyvitamin D3 receptor?

Abstract Highly purified elongation factor 1 (light form, EF1L) from rat liver contains zinc as determined by atomic absorption spectrophotometry. Analysis has been performed on the most active protein fraction from DEAE-Sephadex chromatography (estimated purity: 90%) and on the main band obtained from this fraction by polyacrylamide gel electrophoresis. The data are consistent with a stoichiometry of approximately one g-atom of zinc per 54,000 daltons of EF1L protein. A functional role for Zn2+ is suggested by the fact that 0.3 mM 1,10-phenanthroline completely abolishes GTP binding by EF1L (measured by the nitrocellulose filter retention assay), while the isomeric non-chelator 1,7-phenanthroline has no effect. This inhibition can be overcome by the addition of excess zinc ion.

Synthesis and characterization of a 29-amino acid residue DNA-binding peptide derived from α/β-type small, acid-soluble spore proteins (SASP) of bacteria

Abstract Laser-excited Raman spectra of yeast tRNAs in charged and uncharged states are compared to reveal differences in base stacking. Aminoacylation of unfractionated tRNA reduces considerably the amount of stacking of both adenine and pyrimidine residues. In Phe-tRNA Phe , however, only the adenine residues appear to be significantly less stacked after aminoacylation. The overall degree of order in the backbone of yeast tRNAs is little affected by these changes in base-stacked secondary structure.

Patterns of Mitochondrial DNA Strand Asymmetry Correlate With Phylogeny

BackgroundWe have constructed Bayesian prior-based, amino-acid sequence profiles for the complete yeast mitochondrial proteome and used them to develop methods for identifying and characterizing the context of protein mutations that give rise to human mitochondrial diseases. (Bayesian priors are conditional probabilities that allow the estimation of the likelihood of an event - such as an amino-acid substitution - on the basis of prior occurrences of similar events.) Because these profiles can assemble sets of taxonomically very diverse homologs, they enable identification of the structurally and/or functionally most critical sites in the proteins on the basis of the degree of sequence conservation. These profiles can also find distant homologs with determined three-dimensional structures that aid in the interpretation of effects of missense mutations.ResultsThis survey reports such an analysis for 15 missense mutations, one insertion and three deletions involved in Lebers hereditary optic neuropathy, Leigh syndrome, mitochondrial neurogastrointestinal encephalomyopathy, Mohr-Tranebjaerg syndrome, iron-storage disorders related to Friedreichs ataxia, and hereditary spastic paraplegia. We present structural correlations for seven of the mutations.ConclusionsOf the 19 mutations analyzed, 14 involved changes in very highly conserved parts of the affected proteins. Five out of seven structural correlations provided reasonable explanations for the malfunctions. As additional genetic and structural data become available, this methodology can be extended. It has the potential for assisting in identifying new disease-related genes. Furthermore, profiles with structural homologs can generate mechanistic hypotheses concerning the underlying biochemical processes - and why they break down as a result of the mutations.