Substrate gating confers steroid specificity to estrogen sulfotransferase.

Estrogen sulfotransferase (EST) exhibits a high substrate specificity and catalytic efficiency toward estrogens such as estradiol (E2) but insignificant ability to sulfate hydroxysteroids such as dehydroepiandrosterone (DHEA). To provide the structural basis for this estrogen specificity, we mutated amino acid residues that constitute the substrate-binding site of EST. Among these mutants, only Tyr-81 decreased E2 and increased DHEA sulfotransferase activities. Substitution for Tyr-81 by smaller hydrophobic residues increased K(m(E2)) for E2 activity, whereas the k(cat(E2)) remained relatively constant. The Y81L mutant exhibited the same DHEA activity as wild-type hydroxysteroid sulfotransferase, for which K(m(DHEA)) remained relatively constant, and k(cat(DHEA)) was markedly increased. The side chain of Tyr-81 is directed at the A-ring of the E2 molecule in the substrate-binding pocket of EST, constituting a steric gate with Phe-142 sandwiching E2 from the opposite side. The present mutagenesis study indicates that the 3beta-hydroxyl group of the DHEA molecule is excluded from the catalytic site of EST through steric hindrance of Tyr-81 with the C-19 methyl group of DHEA. Thus, this stricture-like gating caused by steric hindrance appears to be a structural principle for conferring estrogen specificity to EST.     

Chain-shortening of prostaglandin F2 alpha by rat liver peroxisomes

Liver peroxisomes were isolated from di(2-ethylhexyl)phthalate treated rats by isopycnic sucrose gradient centrifugation of a light mitochondrial fraction. Incubation of prostaglandin F2 alpha with purified peroxisomes resulted in conversion into a more polar product(s). In contrast, incubation with mitochondrial fractions and microsomal fractions under the same conditions did not result in any detectable conversion. The polar material obtained from a preparative incubation was purified by high performance liquid chromatography and characterized by radio-gas chromatography and gas chromatography-mass spectrometry. The structure of the polar compound was shown to be 5,7,11-trihydroxy-tetranorprost-9-enoic acid (tetranor-prostaglandin F1 alpha). Prostaglandin F2 alpha was thus chain-shortened by four carbon atoms.     

Patterns of protein synthesis in E. coli: a catalog of the amount of 140 individual proteins at different growth rates.

The amount of 140 individual proteins of E. coli B/r was measured during balanced growth in five different media. The abundance of each protein was determined from its absolute amount in 14C-glucose-minimal medium and a measurement of its relative amount at each growth rate using a double labeling technique. Separation of the proteins was carried out by two-dimensional gel electrophoresis. This catalog of proteins, combined with 50 additional ribosomal proteins already studied, comprises about 5% of the coding capacity of the genome, but accounts for two thirds of the cell's protein mass. The behavior of most of these proteins could be described by a relatively small number of patterns. 102 of the 140 proteins exhibited nearly linear variations with growth rate. The remaining 38 proteins exhibited levels which seemed to depend more on the chemical nature of the medium than on growth rate. Proteins, including the ribosomal proteins, that increase in amount with increasing growth rate account for 20% of total cell protein by weight during growth on acetate, 32% on glucose-minimal medium and 55% on glucose-rich medium. Proteins with invariant levels in the various media comprise about 4% of the cell's total protein.     

QUANTITATIVE CHANGES WITH AGE IN THE DNA CONTENT OF METHYLAZOXYMETHANOL-INDUCED MICROENCEPHALIC RAT BRAIN1

Abstract— DNA synthesis in methylazoxymethanol (MAM)-treated foetal brain was reduced during the first 3 days after the injection of the compound into the mother rat. The MAM-treated brain grew at almost the normal rate after this period, but the reduction in DNA persisted through maturity of the animal. This difference in DNA content between normal and microencephalic brain was restricted to the cerebral hemispheres. The major increase in DNA content of prenatal brain occurred in the cerebrum, whereas the postnatal increase took place in the cerebellum. jH-Labelled MAM was incorporated more extensively into foetal brain DNA than into RNA. The half-life of the MAM-modified nucleic acids was 4–5 days. We suggest that removal of necrotic cells from the brain may account for the rapid loss of label from nucleic acids.     

Structure of casein micelles studied by small-angle neutron scattering

The structure of casein micelles has been studied by small-angle neutron scattering and static light scattering. Alterations in structure upon variation of pH and scattering contrast, as well as after addition of chymosin, were investigated. The experimental data were analyzed by a model in which the casein micelle consists of spherical submicelles. This model gave good agreement with the data and gave an average micellar radius of about 100–120 nm and a submicellar radius of about 7 nm both with a polydispersity of about 40–50%. The contrast variation indicated that the scattering length density of the submicelles was largest at the center of the submicelles. The submicelles were found to be closely packed, the volume fraction varying slightly with pH. Upon addition of chymosin the submicellar structure remained unchanged within the experimental accuracy. Correspondence to: S. Hansen     

Interaction of 8-anilino-l-naphthalene sulfonate with fatty acid treated mycelium of Boletus variegatus Fr.

Summary A fluorescent dye (8-anilino-1-naphthalene-sulphonic acid), which gives no fluorescence, or only a weak one, when added to submerged grown mycelium of the hymenomycete Boletus variegatus, is found to result in a marked increase of fluorescence if the organism is treated with fatty acids. The increased fluorescence is dependent on the presence of undissociated acid molecules as well as on the chain length of the acid. The effect is not due to the acidic character of the molecules, but is highly dependent on lipophilic properties of the substances. Consequently, the fatty acids introduce changes in the characteristics of the cytoplasmic membrane probably as a result of interactions with lipophilic parts of the membrane. The paper finally discusses the contribution of the present results to the elucidation of the influence mechanism of fatty acids in regard to leaking and respiration