The recovery of normal DNA replication kinetics in ultraviolet-irradiated Chinese hamster cells

The kinetics of DNA replication were analyzed in the second S phase following UV irradiation of Chinese hamster ovary cells synchronized at the beginning of S phase. The cells were synchronized by treating cells selected in mitosis with hydroxyurea for 9 h. Following UV irradiation, the cells were allowed to progress until the next mitosis; at which time they were resynchronized at the beginning of the second S phase by the same procedure. The kinetics of DNA replication were determined by measuring the proportion of DNA which achieved hybrid buoyant density on CsCl density gradients as a function of the time of incubation in the presence of 5-bromodeoxyuridine.The results of these experiments showed that even though the rate of DNA replication is substantially depressed during the first S phase following UV irradiation with a fluence of 5 J/m², the rate has recovered to the extent that it is indistinguishable from the unirradiated control by the time the cells have entered their second S phase. It was concluded from these observations that the lesions in DNA which caused the rate of DNA replication to be initially depressed during the first S phase have been either removed or modified such that they no longer are able to cause a reduction in the rate of DNA replication in the second S phase following UV irradiation.     

Disparity in Physician Perception of Patients' Adherence to Medications by Obesity Status

Physician perception of medication adherence may alter prescribing patterns. Perception of patients has been linked to readily observable factors, such as race and age. Obesity shares a similar stigma to these factors in society. We hypothesized that physicians would perceive patients with a higher BMI as nonadherent to medication. Data were collected from the baseline visit of a randomized clinical trial of patient–physician communication (240 patients and 40 physicians). Physician perception of patient medication adherence was measured on a Likert scale and dichotomized as fully adherent or not fully adherent. BMI was the predictor of interest. We performed Poisson regression analyses with robust variance estimates, adjusting for clustering of patients within physicians, to examine the association between BMI and physician perception of medication adherence. The mean (s.d.) BMI was 32.6 (7.7) kg/m². Forty‐five percent of patients were perceived as nonadherent to medications by their physicians. Higher BMI was significantly and negatively associated with being perceived as adherent to medication (prevalence ratio (PrR) 0.76, 95% confidence interval (CI): 0.64–0.90; P = 0.002; per 10 kg/m² increase in BMI). BMI remained significantly and negatively associated with physician perception of medication adherence after adjustment for patient and physician characteristics (PrR 0.80, 95% CI: 0.66–0.96; P = 0.020). In this study, patients with higher BMI were less likely to be perceived as adherent to medications by their providers. Physician perception of medication adherence has been shown to affect prescribing patterns in other studies. More work is needed to understand how this perception may affect the care of patients with obesity.     

NMR spectroscopy of 113Cd(II)-substituted gene 32 protein

Gene 32 protein (g32P), the single-stranded DNA binding protein from bacteriophage T4, contains 1 mol of Zn(II)/mol of protein. This intrinsic zinc is retained within the DNA-binding core fragment, g32P-(A+B) (residues 22-253), obtained by limited proteolysis of the intact protein. Ultraviolet circular dichroism provides evidence that Zn(II) binding causes significant changes in the conformation of the peptide chain coupled with alterations in the microenvironments of tryptophan and tyrosine side chains. NMR spectroscopy of the 113Cd(II) derivative of g32P-(A+B) at both 44.4 and 110.9 MHz shows a single 113Cd resonance, delta 637, a chemical shift consistent with coordination to three of the four sulfhydryl groups in the protein. In vitro mutagenesis of Cys166 to Ser166 creates a mutant g32P that still contains 1 Zn(II)/molecule. This mutant protein when substituted with 113Cd(II) shows a 113Cd signal with a delta and a line width the same as those observed for the wild-type protein. Thus, the S-ligands to the metal ion appear to be contributed by Cys77, Cys87, and Cys90. Relaxation data suggest that chemical shift anisotropy is the dominant, but not exclusive, mechanism of relaxation of the 113Cd nucleus in g32P, since a dipolar modulation from ligand protons is observed at 44.4 MHz but not at 110.9 MHz. Complexation of core 113Cd g32P with d(pA)6 or Co(II) g32P with poly(dT) shows only minor perturbation of the NMR signal or d-d electronic transitions, respectively, suggesting that the metal ion in g32P does not add a ligand from the bound DNA.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of the transciption inhibitory protein, TF1, on phage SP01 promoter complex formation and stability.

The uptake of a homologous single-stranded fragment by superhelical DNA produces a complex that contains a stable displacement loop. When the circular DNA was relaxed by the random action of pancreatic DNAase, complexes dissociated by a process which requires that the single-stranded arm of the D-loop be intact. We attribute the dissociation to branch migration, the exchange of like strands at a branch point. The kinetics of dissociation were biphasic. A fraction of the nicked complexes dissociated in a few seconds, the rest dissociated much more slowly. The fraction of molecules that dissociated slowly was directly related to the length of the third strand, and inversely related to temperature. Salt also inhibited dissociation. Under physiological conditions, 37 °C and 0.15 m-NaCl, more than half of complexes containing a third strand of 1000-nucleotide residues survived for at least one minute. These observations provide a guide to handling certain natural or synthetic branched derivatives of DNA. Analyzing our data by the method of Thompson et al. (1976), we have estimated that the time for the exchange of one nucleotide for another at a single-stranded branch is 12 microseconds; but the calculated value depends strongly upon the assumption that single-strand branch migration occurs by a random walk.     

Molecular basis of biodegradation of chloroaromatic compounds

Chlorinated aromatic hydrocarbons are widely used in industry and agriculture, and comprise the bulk of environmental pollutants. Although simple aromatic compounds are biodegradable by a variety of degradative pathways, their halogenated counterparts are more resistant to bacterial attack and often necessitate evolution of novel pathways. An understanding of such evolutionary processes is essential for developing genetically improved strains capable of mineralizing highly chlorinated compounds. This article provides an overview of the genetic aspects of dissimilation of chloroaromatic compounds and discusses the potential of gene manipulation to promote enhanced evolution of the degradative pathways.     

A SENSITIVE RADIOIMMUNOASSAY FOR 7S NERVE GROWTH FACTOR ANTIGENS IN SERUM AND TISSUES

A radioimmunoassay was developed which can measure accurately concentrations of mouse 7S nerve growth factor antigens (NGFA) as low as 3·0 ng/ml in serum or tissue homogenates. Extremely large amounts of presumed nerve growth factor were found in the submaxillary gland; but considerable quantities were also present in mouse serum, kidney, adrenal gland and vas deferens. Heart, spleen, liver and muscle contained less of the presumed nerve growth factor, and only small amounts were recovered from brain. Rat adrenal gland and serum from rats, guinea pigs and man contained much less immunologically reactive material. The level of presumed nerve growth factor in the mouse heart was highest at birth and decreased slowly during maturation. In the mouse submaxillary gland the content of presumed nerve growth factor increased rapidly after 2 weeks of postnatal age, with higher levels found in male animals. Destruction with 6-hydroxydopamine of the sympathetic nerves in the hearts of newborn or adult mice did not significantly alter the amount of presumed nerve growth factor recovered in the heart.