Unusual features of the c-ring of F1FO ATP synthases.

Membrane integral ATP synthases produce adenosine triphosphate, the universal "energy currency" of most organisms. However, important details of proton driven energy conversion are still unknown. We present the first high-resolution structure (2.3 Å) of the in meso crystallized c-ring of 14 subunits from spinach chloroplasts. The structure reveals molecular mechanisms of intersubunit contacts in the c14-ring, and it shows additional electron densities inside the c-ring which form circles parallel to the membrane plane. Similar densities were found in all known high-resolution structures of c-rings of F1FO ATP synthases from archaea and bacteria to eukaryotes. The densities might originate from isoprenoid quinones (such as coenzyme Q in mitochondria and plastoquinone in chloroplasts) that is consistent with differential UV-Vis spectroscopy of the c-ring samples, unusually large distance between polar/apolar interfaces inside the c-ring and universality among different species. Although additional experiments are required to verify this hypothesis, coenzyme Q and its analogues known as electron carriers of bioenergetic chains may be universal cofactors of ATP synthases, stabilizing c-ring and prevent ion leakage through it.

New Insights on Signal Propagation by Sensory Rhodopsin II/Transducer Complex.

The complex of two membrane proteins, sensory rhodopsin II (NpSRII) with its cognate transducer (NpHtrII), mediates negative phototaxis in halobacteria N. pharaonis. Upon light activation NpSRII triggers a signal transduction chain homologous to the two-component system in eubacterial chemotaxis. Here we report on crystal structures of the ground and active M-state of the complex in the space group I212121. We demonstrate that the relative orientation of symmetrical parts of the dimer is parallel ("U"-shaped) contrary to the gusset-like ("V"-shaped) form of the previously reported structures of the NpSRII/NpHtrII complex in the space group P21212, although the structures of the monomers taken individually are nearly the same. Computer modeling of the HAMP domain in the obtained "V"- and "U"-shaped structures revealed that only the "U"-shaped conformation allows for tight interactions of the receptor with the HAMP domain. This is in line with existing data and supports biological relevance of the "U" shape in the ground state. We suggest that the "V"-shaped structure may correspond to the active state of the complex and transition from the "U" to the "V"-shape of the receptor-transducer complex can be involved in signal transduction from the receptor to the signaling domain of NpHtrII.

High-resolution structure of a membrane protein transferred from amphipol to a lipidic mesophase.

Amphipols (APols) have become important tools for the stabilization, folding, and in vitro structural and functional studies of membrane proteins (MPs). Direct crystallization of MPs solubilized in APols would be of high importance for structural biology. However, despite considerable efforts, it is still not clear whether MP/APol complexes can form well-ordered crystals suitable for X-ray crystallography. In the present work, we show that an APol-trapped MP can be crystallized in meso. Bacteriorhodopsin (BR) trapped by APol A8-35 was mixed with a lipidic mesophase, and crystallization was induced by adding a precipitant. The crystals diffract beyond 2 Å. The structure of BR was solved to 2 Å and found to be indistinguishable from previous structures obtained after transfer from detergent solutions. We suggest the proposed protocol of in meso crystallization to be generally applicable to APol-trapped MPs.

Nanoparticle surface-enhanced Raman scattering of bacteriorhodopsin stabilized by amphipol A8-35.

Surface-enhanced Raman spectroscopy (SERS) has developed dramatically since its discovery in the 1970s, because of its power as an analytical tool for selective sensing of molecules adsorbed onto noble metal nanoparticles (NPs) and nanostructures, including at the single-molecule (SM) level. Despite the high importance of membrane proteins (MPs), SERS application to MPs has not really been studied, due to the great handling difficulties resulting from the amphiphilic nature of MPs. The ability of amphipols (APols) to trap MPs and keep them soluble, stable, and functional opens up onto highly interesting applications for SERS studies, possibly at the SM level. This seems to be feasible since single APol-trapped MPs can fit into gaps between noble metal NPs, or in other gap-containing SERS substrates, whereby the enhancement of Raman scattering signal may be sufficient for SM sensitivity. The goal of the present study is to give a proof of concept of SERS with APol-stabilized MPs, using bacteriorhodopsin (BR) as a model. BR trapped by APol A8-35 remains functional even after partial drying at a low humidity. A dried mixture of silver Lee-Meisel colloid NPs and BR/A8-35 complexes give rise to SERS with an average enhancement factor in excess of 10(2). SERS spectra resemble non-SERS spectra of a dried sample of BR/APol complexes.

Ground state structure of D75N mutant of sensory rhodopsin II in complex with its cognate transducer.

The complex of sensory rhodopsin II (NpSRII) with its cognate transducer (NpHtrII) mediates negative phototaxis in halobacteria Natronomonas pharaonis. Upon light activation NpSRII triggers, by means of NpHtrII, a signal transduction chain homologous to the two component system in eubacterial chemotaxis. Here we report on the crystal structure of the ground state of the mutant NpSRII-D75N/NpHtrII complex in the space group I212121. Mutations of this aspartic acid in light-driven proton pumps dramatically modify or/and inhibit protein functions. However, in vivo studies show that the similar D75N mutation retains functionality of the NpSRII/NpHtrII complex. The structure provides the molecular basis for the explanation of the unexpected observation that the wild and the mutant complexes display identical physiological response on light excitation.

Finasteride in the treatment of men with frontal male pattern hair loss.

BACKGROUND: Finasteride, a specific inhibitor of type II 5alpha-reductase, decreases serum and scalp dihydrotestosterone and has been shown to be effective in men with vertex male pattern hair loss. OBJECTIVE: This study evaluated the efficacy of finasteride 1 mg/day in men with frontal (anterior/mid) scalp hair thinning. METHODS: This was a 1-year, double-blind, placebo-controlled study followed by a 1-year open extension. Efficacy was assessed by hair counts (1 cm2 circular area), patient and investigator assessments, and global photographic review. RESULTS: There was a significant increase in hair count in the frontal scalp of finasteride-treated patients (P < .001), as well as significant improvements in patient, investigator, and global photographic assessments. Efficacy was maintained or improved throughout the second year of the study. Finasteride was generally well tolerated. CONCLUSION: In men with hair loss in the anterior/mid area of the scalp, finasteride 1 mg/day slowed hair loss and increased hair growth.

Arabidopsis thaliana centromere regions: genetic map positions and repetitive DNA structure.

The genetic positions of the five Arabidopsis thaliana centromere regions have been identified by mapping size polymorphisms in the centromeric 180-bp repeat arrays. Structural and genetic analysis indicates that 180-bp repeat arrays of up to 1000 kb are found in the centromere region of each chromosome. The genetic behavior of the centromeric arrays suggests that recombination within the arrays is suppressed. These results indicate that the centromere regions of A. thaliana resemble human centromeres in size and genomic organization.

Clinical experience of the detection of prostate cancer in patients with benign prostate hyperplasia treated with finasteride. The Finasteride Study Group.

The clinical experience relating to the detection of prostate cancer in patients participating in 2 large multicenter clinical trials of finasteride in the treatment of benign prostatic hyperplasia is reviewed. A total of 1,645 patients 40 to 83 years old with benign prostatic hyperplasia was randomized to receive 1 or 5 mg finasteride or placebo once a day for 12 months in a double-blind fashion followed by an open extension study in which all patients were treated with 5 mg finasteride daily. At entry, all patients were to have a maximum urinary flow rate of 15 ml per second or less with a voided volume of 150 ml or more, an enlarged prostate and symptoms of urinary obstruction. Patients with a prostate specific antigen level of 40 ng/ml or more, or any finding suggestive of prostate cancer were excluded. During the study period 32 cases of prostate cancer were diagnosed: 12 were detected during the 12 months of the controlled study and were evenly distributed among the treatment groups (4 on placebo, and 3 on 1 mg and 5 on 5 mg finasteride) and 20 cases were detected in the extension study. From these results we conclude that finasteride-treated patients should be evaluated periodically by digital rectal examination, careful monitoring of prostate specific antigen levels and appropriate investigation of any suspicious findings.

Scandinavian clinical study of finasteride in the treatment of benign prostatic hyperplasia.

The effects of finasteride, a potent 5 alpha-reductase inhibitor, were assessed in patients with benign prostatic hyperplasia. Patients were treated with finasteride or placebo for 24 weeks in a double-blind multicenter study followed by a 12-month open-extension period. After 24 weeks, finasteride-treated patients, when compared to placebo-treated patients, showed a significant reduction in prostate volume (22.5% median decrease) and prostate significant antigen (32.4% median decrease), a significant increase in maximum urinary flow (1.6 ml/s mean increase from baseline) and a significant improvement in their obstructive symptom scores (two-point decrease from baseline). Finasteride was well tolerated, and the improvements in prostate volume, maximum urinary flow rate and obstructive symptom scores observed in the controlled study were maintained throughout the extension study.

Norfloxacin, amoxycillin, cotrimoxazole and nalidixic acid. A summary of 3-day and 7-day therapy studies in the treatment of urinary tract infections.

The results of clinical trials in which norfloxacin was used for 7 days compared with amoxycillin or cotrimoxazole, or for 3 days compared with citrated nalidixic acid, are presented. Additionally, the results of a concurrent open study of 3 days of norfloxacin in the management of simple urinary tract infections are discussed. Resistance to norfloxacin was only encountered in 0.2% of pathogens isolated. Norfloxacin was as effective in eradicating bacteriuria as amoxycillin, cotrimoxazole or citrated nalidixic acid. The response to 3 days of norfloxacin was similar to that seen after 7 days therapy with this compound, or to 7 days of cotrimoxazole. The incidence of adverse experiences to norfloxacin in 758 patients was below 10%.

Norfloxacin versus cotrimoxazole in the treatment of uncomplicated urinary tract infections--a multi-centre trial.

One hundred and twenty-two patients with uncomplicated urinary tract infections were treated with either 400 mg bd norfloxacin or 160/800 mg bd cotrimoxazole for 7 days. Follow-up examinations showed norfloxacin to be equally effective as cotrimoxazole in the eradication of bacteriuria and symptoms. Norfloxacin caused fewer and less severe side effects.

The inadequacy of rectal temperature measurements for assessing the effects of antiviral drugs on influenza virus infection of ferrets.

From 6 experiments in which 99 ferrets were infected with influenza virus A/Finland/74 and treated with various agents which suppress virus shedding and other parameters of infection, we assessed whether rectal temperature correlated with nasal virus shedding. A number of temperature and virus-shedding related parameters were determined for each experiment but statistical analysis showed little correlation between them, although an elevated temperature occurred at some time after infection. The pooled data also suggested that temperature and virus shedding parameters are not clearly related. The analysis indicates that intermittent rectal temperature measurements are unsatisfactory for determining the efficacy of anti-influenza agents in ferrets.

Antiviral effects of single-stranded polynucleotide inhibitors of the influenza virion-associated transcriptase against influenza virus infection of hamsters and ferrets.

Administration of a single-stranded polynucleotide copolymer containing 9% cytidine residues and 91% 4-thiouridine residues [poly(C,S4U10)], a known potent inhibitor of the virion transcriptase of influenza viruses, suppressed the amount of virus recoverable from the nasal washes of influenza virus-infected hamsters and ferrets. The incidence of sneezing and nasal discharge in infected ferrets was also reduced. In hamsters, poly(C,S4U10) was more effective than amantadine-HCl or Virazole. Polyinosinic acid in combination with poly-5-hydroxy cytidylic acid also had anti-influenza effects. Poly(C,S4U10) annealed to polyadenylic acid was not effective, nor was the double-stranded polymer (polyinosinic acid) . (polycytidylic acid) even when complexed with carboxymethylcellulose and polylysine. No toxic effects of poly(C,S4U10) were apparent in the treated hamsters and ferrets, and high doses (greater than or equal to 2.86 g/kg) administered intraperitoneally to mice produced no adverse effects.

A putative interferon induced in hamsters by poly(I) . poly(C).

Encephalomyocarditis (EMC) virus causes lethal infection of hamsters against which poly(I) . poly(C) causes dose-dependent protection. In contrast, no antiviral effects occur with poly(I) . poly(C) against influenza virus infection of hamsters. Serum from poly(I) . poly(C) treated hamsters protects other hamsters against EMC virus infection with maximum protection with serum removed 3h after poly(I) . poly(C) treatment of the donor hamsters. In such assays the factor was found to be inactivated by trypsin and pH 2 and 56 degrees C for 1 hr. The serum factor did not confer protection against EMC virus infection of L-929, BHK, Hak or primary hamster embryo cells. The amount of poly(I) . poly(C) carried over into serum samples of poly(I) . poly(C) treated hamsters was insufficient to account for the antiviral effects. The antiviral serum factor is presumed to be a form of interferon despite the fact that it does not titrate in cell cultures and has a novel set of properties from those which describe known interferons.