Expression of enzymatically active reverse transcriptase in Escherichia coli

Reverse transcriptase of murine retroviruses is a monomeric protein of approximately 80,000 daltons, which is encoded by the central portion of the viral pol gene. To prepare large quantities of the enzyme, we have constructed gene fusions between the trpE gene and portions of the pol gene of Moloney murine leukemia virus. The inserted pol gene sequences include the entire coding region for the mature enzyme and various amounts of additional coding sequences. Many of these constructs express high levels of reverse transcriptase activity even though the NH2 and COOH termini of the protein product only approximate the correct termini of the authentic protein

Policy Formulation Versus Policy Implementation Under the Magnuson-Stevens Fishery Conservation and Management Act: Insight from the North Pacific Crab Rationalization

The Magnuson-Stevens Fishery Conservation and Management Act (MSA) governs management of fisheries located three to 200 miles off the coast of the United States. The MSA is unique in administrative law in that it devolves policy formulation to eight Regional Fishery Management Councils rather than to a federal agency. That agency, the National Marine Fisheries Service (NMFS), is relegated primarily to developing regulations that implement the councils’ policies. NMFS can review the councils’ policies only to ensure that they are consistent with existing laws. NMFS has no authority to revise policy to suit its own preferences, or to write regulations that undercut council policy intent, except when conflicts with other applicable laws arise. The MSA’s legislative history reveals NMFS routinely undercuts this special administrative process through the regulations it writes. We review a recent example in which NMFS attempted to undermine the North Pacific Fishery Management Council’s crab rationalization policy through the regulation-writing process. We offer a simple solution to help avoid future abuse of administrative authority. This solution may have utility in other areas of administrative law in which authority to formulate policy is separated from the power to implement it

Near-infrared surface photometry of early-type galaxies

CCD infrared (JHKs) photometry was performed on a sample of 10 elliptical and 2 lenticular galaxies. Isophotal parameters, brightness profiles, integrated colors and color gradients arepresented. Color gradients found arevery weak, showing bluer colors towards the outer regions. The colors of the sample galaxies are compatible with stellar populations like those found in metal-rich clusters of the Galaxy; objects NGC7192, NGC7562 and NGC7619 are compatible with less metal-rich populations. The brightness profile of most galaxies is well described by the r1/4 law. The profiles of NGC1600 and NGC720 are described by Sérsic’s law with n ∼ 1.5 andn ∼ 1.8 respectively. The infrared effective radius of the objects studied is typically one half of its counterpart in the B band, which can be an indication that the stellar population that dominates the infrared emission is more concentrated in the central regions. Weshow that the sample satisfies the Fundamental Plane relation of elliptical galaxies in the infrared, with an rms scatter of 0.20 for J and H and 0.23 for Ks

Differential multimerization of Moloney murine leukemia virus integrase purified under nondenaturing conditions

Retroviral integrases (IN) catalyze the integration of the reverse-transcribed viral DNA into the host genome, an essential process leading to virus replication. For Moloney murine leukemia virus (M-MuLV) IN, the limited solubility of the recombinant protein has restricted the development of biophysical and structural analyses. Herein, recombinant M-MuLV IN proteins, either full length or two nonoverlapping domain constructs, were purified under non-denaturing conditions from solubilized bacterial extracts by Ni2+-NTA resins. Additionally, WT IN was further purified by heparin chromatography. All of the purified proteins were shown to be active and stable. WT M-MuLV IN chromatographed with a peak corresponding with a dimer by gel filtration chromatography. In contrast, the single point mutant C209A IN migrated predominantly as a tetramer. For both proteins, fractions in equilibrium between dimers and tetramers were competent to assemble concerted two-end integrations and yielded a unique strand-transfer profile in the presence of a 28-mer U5 oligonucleotide substrate, indicative of a distinct conformation within the synaptic complex. This specific target-site selection was not observed with a shorter 20-mer U5 substrate. These studies provide the foundation for biophysical and structural analysis on M-MuLV IN and the mechanism of retroviral integration

Socialist Dandies International: East Europe, 1946-1959

This article maps the looks and lifestyle choices of small groups of young, like-minded people who emerged in the postwar Soviet Union and East Europe in the background of huge political, social, and cultural changes. With their androgynous bodies wrapped in drape jackets and narrow trousers, and their love of jazz and swing, these young men stood in a sharp contrast to the official ideology that promoted socialism as a new, pure, and highly rationalized project, its ideal robust and strong man, and its mass culture that insisted on educational and restrained forms of entertainment. Through the categories of dress, body, and big city, the article investigates the clashes, and the eventual truce, between the socialist streamlined and rationalized master narrative and the young dandies' fragmented and disordered narrative. The article argues that the socialist dandies were not politically minded, and that their challenge to the officially proclaimed values was informed by their adolescent recklessness and a general postwar desolation. They were declared state enemies because the socialist regimes did not allow for alternative types of modernity. Consequently, the authorities condemned the young dandies' looks and interests as cosmopolitan, because they originated in the West, and as artificial, since they belonged to the culture that had preceded a new socialist world

Is Hip Abduction Strength Asymmetry Present in Female Runners in the Early Stages of Patellofemoral Pain Syndrome?

BACKGROUND: The current literature indicates that hip abduction weakness in female patients is associated with ipsilateral patellofemoral pain syndrome (PFPS) as part of the weaker hip abductor complex. Thus, it has been suggested that clinicians should consider screening female athletes for hip strength asymmetry to identify those at risk of developing PFPS to prevent the condition. However, no study to date has demonstrated that hip strength asymmetry exists in the early stages of PFPS. PURPOSE: To determine whether hip abduction strength asymmetry exists in female runners with early unilateral PFPS, defined as symptoms of PFPS not significant enough to cause patients to seek medical attention or prevent them from running at least 10 miles per week. STUDY DESIGN: Controlled laboratory study. METHODS: This study consisted of 21 female runners (mean age, 30.5 years; range, 18-45 years) with early unilateral PFPS, who had not yet sought medical care and who were able to run at least 10 miles per week, and 36 healthy controls comparably balanced for age, height, weight, and weekly running mileage (mean, 18.5 mi/wk). Study volunteers were recruited using flyers and from various local running events in the metropolitan area. Bilateral hip abduction strength in both a neutral and extended hip position was measured using a handheld dynamometer in each participant by an examiner blinded to group assignment. RESULTS: Patients with early unilateral PFPS demonstrated no significant side-to-side difference in hip abduction strength, according to the Hip Strength Asymmetry Index, in both a neutral (mean, 83.5 ± 10.2; P = .2272) and extended hip position (mean, 96.3 ± 21.9; P = .6671) compared with controls (mean, 87.0 ± 8.3 [P = .2272] and 96.6 ± 16.2 [P = .6671], respectively). Hip abduction strength of the affected limb in patients with early unilateral PFPS (mean, 9.9 ± 2.2; P = .0305) was significantly stronger than that of the weaker limb of control participants (mean, 8.9 ± 1.4; P = .0305) when testing strength in a neutral hip position; however, no significant difference was found when testing the hip in an extended position (mean, 7.0 ± 1.4 [P = .1406] and 6.6 ± 1.5 [P =.1406], respectively). CONCLUSION: The study data show that early stages of unilateral PFPS in female runners is not associated with hip abduction strength asymmetry and that hip abduction strength tested in neutral is significantly greater in the affected limb in the early stages of PFPS compared with the unaffected limb. However, when tested in extension, no difference exists. Further studies investigating the early stages of PFPS are warranted. CLINICAL RELEVANCE: Unlike patients with PFPS seeking medical care, early PFPS does not appear to be significantly associated with hip abduction strength asymmetry

Increased Matrix Metalloproteinase (MMPs) Levels Do Not Predict Disease Severity or Progression in Emphysema

Rationale: Though matrix metalloproteinases (MMPs) are critical in the pathogenesis of COPD, their utility as a disease biomarker remains uncertain. This study aimed to determine whether bronchoalveolar lavage (BALF) or plasma MMP measurements correlated with disease severity or functional decline in emphysema. Methods: Enzyme-linked immunosorbent assay and luminex assays measured MMP-1, -9, -12 and tissue inhibitor of matrix metalloproteinase-1 in the BALF and plasma of non-smokers, smokers with normal lung function and moderate-to-severe emphysema subjects. In the cohort of 101 emphysema subjects correlative analyses were done to determine if MMP or TIMP-1 levels were associated with key disease parameters or change in lung function over an 18-month time period. Main Results: Compared to non-smoking controls, MMP and TIMP-1 BALF levels were significantly elevated in the emphysema cohort. Though MMP-1 was elevated in both the normal smoker and emphysema groups, collagenase activity was only increased in the emphysema subjects. In contrast to BALF, plasma MMP-9 and TIMP-1 levels were actually decreased in the emphysema cohort compared to the control groups. Both in the BALF and plasma, MMP and TIMP-1 measurements in the emphysema subjects did not correlate with important disease parameters and were not predictive of subsequent functional decline. Conclusions: MMPs are altered in the BALF and plasma of emphysema; however, the changes in MMPs correlate poorly with parameters of disease intensity or progression. Though MMPs are pivotal in the pathogenesis of COPD, these findings suggest that measuring MMPs will have limited utility as a prognostic marker in this disease. © 2013 D'Armiento et al

Inhibition of Moloney murine leukemia virus integration using polyamides targeting the long-terminal repeat sequences

The retroviral integrase (IN) carries out the integration of the viral DNA into the host genome. Both IN and the DNA sequences at the viral long-terminal repeat (LTR) are required for the integration function. In this report, a series of minor groove binding hairpin polyamides targeting sequences within terminal inverted repeats of the Moloney murine leukemia virus (M-MuLV) LTR were synthesized, and their effects on integration were analyzed. Using cell-free in vitro integration assays, polyamides targeting the conserved CA dinucleotide with cognate sites closest to the terminal base pairs were effective at blocking 3' processing but not strand transfer. Polyamides which efficiently inhibited 3' processing and strand transfer targeted the LTR sequences through position 9. Polyamides that inhibited integration were effective at nanomolar concentrations and showed subnanomolar affinity for their cognate LTR sites. These studies highlight the role of minor groove interactions within the LTR termini for retroviral integration

Inhibition of Moloney murine leukemia virus integration using polyamides targeting the long-terminal repeat sequences

The retroviral integrase (IN) carries out the integration of the viral DNA into the host genome. Both IN and the DNA sequences at the viral long-terminal repeat (LTR) are required for the integration function. In this report, a series of minor groove binding hairpin polyamides targeting sequences within terminal inverted repeats of the Moloney murine leukemia virus (M-MuLV) LTR were synthesized, and their effects on integration were analyzed. Using cell-free in vitro integration assays, polyamides targeting the conserved CA dinucleotide with cognate sites closest to the terminal base pairs were effective at blocking 3' processing but not strand transfer. Polyamides which efficiently inhibited 3' processing and strand transfer targeted the LTR sequences through position 9. Polyamides that inhibited integration were effective at nanomolar concentrations and showed subnanomolar affinity for their cognate LTR sites. These studies highlight the role of minor groove interactions within the LTR termini for retroviral integration