Cloning of nisin resistance determinant and replication origin on 7.6-kilobase EcoRI fragment of pNP40 from Streptococcus lactis subsp. diacetylactis DRC3

The nisin resistance determinant and an origin of replication on pNP40, a plasmid of about 60 kilobases that is present in Streptococcus lactis subsp. diacetylactis DRC3, was cloned on a 7.6-kilobase EcoRI fragment. When self-ligated, this fragment existed as an independent replicon (pFM011) and contained a 2.6-kilobase EcoRI-XbaI fragment encoding nisin resistance.     

Structure determination of the 2:1 derivatives of copper(I) bromide and iodide with bis(diphenylphosphino)methane. A simple structural scheme for the formation of (CuX)nLm species

The structures of the complexes (CuX)₂DPM (X = Br, I; DMP = bis(diphenylphosphino)methane) were determined from three dimensional X-ray data collected by counter methods. The iodine derivative crystallizes in the space group Pbca with eight units in a cell defined by a = 17.128(9), b = 18.306(9) c, = 16.508 (8) Å. The structure was refined by the least-squares method to a final R factor of 0.054 for 1336 non-zero independent reflections. The bromine derivative crystallizes in the space group P2₁/c with eight units in a cell defined by a = 23.707(1), b = 17.805(9), c = 16.991(1) Å, β = 136.10(5)°. The final least-squares refinement, based on 2489 non-zero independent reflections, gave an R factor of 0.074.Both the compounds have similar structures with a centrosymmetric (CuX)₄ core, in which two copper atoms have a tetrahedral geometry, while the other two are trigonal.The above structures are compared with those already reported for other compounds (CuX)_nL_m and a single scheme is proposed to rationalize the different geometries of the (CuX)_n core on the basis of steric and electronic effects.     

Turnover of hepatic phosphofructokinase in normal and diabetic rats. Role of insulin and peptide stabilizing factor.

Earlier work demonstrated that the activity of liver phosphofructokinase (PFK-L2) and immunoreactive PFK-L2 were decreased in diabetic rats and increased to normal or super-normal amounts following insulin treatment (Dunaway, G.A., and Weber, G., (1974) Arch. Biochem. Biophys. 162, 629-637). This report indicates that the decrease in levels of PFK-L2 in diabetic rats is a result of an accelerated degradation rate while the synthetic rate remains nearly normal. Following insulin treatment, the rate of PFK-L2 synthesis is enhanced 2-fold, whereas the rate of degradation appears to be greatly diminished. An inverse relationship is shown to exist between the PFK-L2 levels and the rates of PFK-L2 degradation, suggesting that the levels of PFK-L2 are primarily regulated by degradation rate. In addition, the levels of the PFK-L2 peptide stabilizing factor are inversely proportional to rates of PFK-L2 degradation. These results indicate that insulin mediates the rate of degradation of PFK-L2 by controlling the level of the peptide stabilizing factor.     

Calcium-dependent binding of EDTA-extractable proteins to calf lens fiber membrane structures

Calf lens fiber membranes and fractions enriched in junction-like structures have been isolated in the absence and presence of EDTA. Their biochemical features have been studied. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and immunoblotting experiments have provided evidence that a distinct group of EDTA-extractable proteins, being one of the main protein components of calf lens fiber membranes and very likely also of junction-like structures, is bound to these membranes via calcium ions. In addition to these proteins, four polypeptides with apparent molecular weights between 14 000 and 17 000 are characteristic for detergent-insoluble lens fiber structures prepared in calcium-rich medium. The absence of EDTA-extractable proteins in the urea-soluble calcium-containing fraction implies that they are not components of the cytoskeleton and that the calcium-dependent binding of these proteins to the membrane is urea-resistant. The use of EDTA throughout the whole membrane isolation procedure results in their complete removal from the membranes which already starts during buffer washing. This indicates that EDTA-extractable proteins exclusively consist of extrinsic membrane proteins which probably are not involved in cytoskeleton binding.     

Effects of subcutaneous injection and intrahypothalamic infusion of releasing hormones upon lordotic response to repetitive coital stimulation.

The effects of luteinizing hormone-releasing hormone (LRH) and thyrotropin-releasing hormone (TRH) upon the lordotic response to repetitive coital stimlation were studied using ovariectomized (OVX) and ovariectomized-adrenalectomized (OVX-ADX) female rats. Both OVX and OVX-ADX rats, pretreated with estrone alone, exhibited a dual behavioral response to repeated coital stimulation. The initial response to short-term stimulation was facilitatory with peak sexual receptivity occurring approximately 120 min following the initial male contact. This initial phase was followed by a depression of sexual receptivity associated with continued coital stimulation. Subcutaneous injection of 500 ng of LRH prior to mating was found to significantly potentiate the initial increases in sexual receptivity and to delay the onset of behavioral depression. The injection of 500 ng of TRH was observed to significantly depress behavioral enhancement due to repetitive coital stimulation.The repetitive coital stimulation model was utilized to localize forebrain areas behaviorally responsive to LRH and TRH. Stainless steel cannulas were implanted into either the medial preoptic area (MPOA), arcuate area (ARC), lateral hypothalamic area (LHA), or cerebral cortex (CC). Cannulated animals, primed with estrone, were tested for sexual receptivity immediately prior to experimental treatment, i.e., the infusion of 0.5 μl of 50 ng of LRH or TRH in 0.9% saline, 0.5 μl of 0.9% saline, or sham infusion. A second mating (postinfusion) test was performed 1.75 hr following infusion. When infused into the MPOA or ARC, LRH significantly enhanced lordotic behavior as compared to values obtained for saline or sham infusions. The infusion of LRH into LHA or CC showed no enhancement beyond the levels observed in control infusions (saline and sham infusions). The infusion of TRH into the MPOA or ARC depressed lordotic enhancement to repeated mating, however, this depression was significant only in ARC. These findings were consistent with previously demonstrated actions of releasing hormones upon neural activity within the MPOA and ARC.