RMCgui: a new interface for the workflow associated with running Reverse Monte Carlo simulations.

The Reverse Monte Carlo method enables construction and refinement of large atomic models of materials that are tuned to give best agreement with experimental data such as neutron and x-ray total scattering data, capturing both the average structure and fluctuations. The practical drawback with the current implementations of this approach is the relatively complex workflow required, from setting up the configuration and simulation details through to checking the final outputs and analysing the resultant configurations. In order to make this workflow more accessible to users, we have developed an end-to-end workflow wrapped within a graphical user interface-RMCgui-designed to make the Reverse Monte Carlo more widely accessible.

Detecting low-level flexibility using residual dipolar couplings: a study of the conformation of cellobiose.

We have developed novel NMR methods for the measurement of heteronuclear residual dipolar couplings (RDCs) in molecules with severely overlapping NMR resonances. These and other methods enabled us to obtain 31 RDCs for α-D-cellobiose and 24 RDCs for β-D-cellobiose. The interpretation of the data in the approximation of a rigid disaccharide structure, using RDCs and interglycosidic (3)J coupling constants, yielded conformation that is very close to that determined using X-ray crystallography.

Preclinical assessment of the efficacy of mycograb, a human recombinant antibody against fungal HSP90.

Mycograb (NeuTec Pharma plc) is a human genetically recombinant antibody against fungal heat shock protein 90 (HSP90). Antibody to HSP90 is closely associated with recovery in patients with invasive candidiasis who are receiving amphotericin B (AMB). Using in vitro assays developed for efficacy assessment of chemotherapeutic antifungal drugs, Mycograb showed activity against a wide range of yeast species (MICs against Candida albicans [fluconazole [FLC]-sensitive and FLC-resistant strains], Candida krusei, Candida tropicalis, Candida glabrata, and Candida parapsilosis, 128 to 256 microg/ml).

Identification of ABC transporters in vancomycin-resistant Enterococcus faecium as potential targets for antibody therapy.

The occurrence of an outbreak of septicaemias due to vancomycin-resistant Enterococcus faecium (VRE), in Manchester, UK, provided an opportunity to examine the antibody responses in patients infected by the same strain. Immunoblotting sera from 24 cases, six of whom died, showed an immunodominant cluster of antigens at 34, 54 and 97 kDa, with a statistically significant correlate between survival and immunoglobulin G to the 34 and 97 kDa bands (P<0.05).

Identification of an immunodominant drug efflux pump in Burkholderia cepacia.

Burkholderia cepacia, a major pathogen amongst individuals with cystic fibrosis (CF), is intrinsically resistant to most clinically available antibiotics. We report the identification of an immunodominant antigen in CF patients infected with B. cepacia, a multidrug-resistance efflux pump called BcrA.

Quantification of Escherichia coli genomic DNA contamination in recombinant protein preparations by polymerase chain reaction and affinity-based collection.

This study describes the development of a novel assay for the quantification of Escherichia coli genomic DNA contamination in recombinant protein samples. The technique is based on PCR amplification and digoxygenin labeling of the genes encoding 5S ribosomal RNA followed by affinity-based collection and detection. Samples containing 1 pg x mL(-1) of extracted E.

The efficacy of acquired humoral and cellular immunity in the prevention and therapy of experimental fungal infections.

In the past two decades, numerous studies have documented the importance of acquired immunity for host defense against invasive fungal infections. There is widespread consensus in the field of medical mycology that cellular immunity is critical for successful host defense against fungi. However, in recent years several studies have established the potential efficacy of humoral immunity in host protection against two major fungal pathogens: Candida albicans and Cryptococcus neoformans.

The molecular cloning, nucleotide sequence and expression of an antigenic determinant from Porphyromonas gingivalis.

A genomic library generated in Escherichia coli was probed with a monoclonal antibody (mAb) LDS28, which reacts with a species-specific cell-surface antigen of Porphyromonas gingivalis. A clone designated pGPR2.1 was shown to express a 46-kDa protein reactive with mAb LDS28, which maps to a 1.

The Bpel locus encodes type III secretion machinery in Bordetella pertussis.

Type III secretory genes(Bscl, J, K, L, N and O) have recently been identified in Bordetella bronchiseptica and shown to be under the control of the BvgAS locus. We examined a 35 616 byte DNA sequence amplified from Bordetella pertussis Tohama I for homology with known type III secretory genes in Yersinia spp. and Pseudomonas sppand a total of 20 homologous open reading frames were detected.

The localization of KpsC, S and T, and KfiA, C and D proteins involved in the biosynthesis of the Escherichia coli K5 capsular polysaccharide: evidence for a membrane-bound complex.

Biosynthesis of the Escherichia coli K5 polysaccharide requires the KfiA, KfiB, KfiC and KfiD proteins. The subsequent transport of the polysaccharide onto the cell surface requires the KpsC, KpsD, KpsE, KpsM, KpsS and KpsT proteins, which are conserved between different group II capsular polysaccharides. The KfiA and KfiC, together with the KpsC, KpsS and KpsT proteins, were purified and polyclonal antisera to each protein generated.

Region 2 of the Escherichia coli K5 capsule gene cluster encoding proteins for the biosynthesis of the K5 polysaccharide.

The nucleotide sequence of region 2 of the Escherichia coli K5 capsule gene cluster has been determined. This region, essential for the biosynthesis of the K5 polysaccharide, contained four genes, termed kfiA-D. The G + C ratio was 33.

Expression and characterization of UDPGlc dehydrogenase (KfiD), which is encoded in the type-specific region 2 of the Escherichia coli K5 capsule genes.

Region 2 of the Escherichia coli K5 capsule gene cluster contains four genes (kfiA through -D) which encode proteins involved in the synthesis of the K5 polysaccharide. A DNA fragment containing kfiD was amplified by PCR and cloned into the gene fusion vector pGEX-2T to generate a GST-KfiD fusion protein. The fusion protein was isolated from the cytoplasms of IPTG (isopropyl-beta-D-thiogalactopyranoside)-induced recombinant bacteria by affinity chromatography and cleaved with thrombin.

Generation and characterization of a monoclonal antibody specific for the major thiol-activated cysteine proteinase of Porphyromonas gingivalis W83.

An IgM monoclonal antibody specified for the thiol-activated proteinase of the oral pathogen Porphyromonas gingivalis W83 was generated. The antibody reacted with a single protein of approximate molecular mass 43 kDa in outer membrane preparations of P. gingivalis.

Cloning and expression of a Porphyromonas (Bacteroides) gingivalis protease gene in Escherichia coli.

A genomic library of Bacteroides gingivalis W83 chromosomal DNA was constructed in the Escherichia coli lambda (lambda) vector EMBL 4. Three recombinant lambda phages expressing a cloned protease were identified in the library. All three lambda phages contained cloned overlapping DNA fragments from the same region of the chromosome and encoded the same cloned protease.

Serum testosterone measurements.

Serum testosterone and especially free testosterone is one of the parameters commonly used to evaluate androgen excess or deficiency. The authors equilibrated serum samples with 14C-labeled testosterone followed by an ammonium sulfate precipitation to compare the "apparent free testosterone concentration" with "total" serum testosterone concentration in the following populations: normal males and females; females presenting with gynecologic problems, particularly hirsutism and/or virilization; and males and females on maintenance hemodialysis. Total serum testosterone for each specimen was assayed with five different commercially available RIA kits encompassing a variety of technics: direct assay technics, assays utilizing extraction procedures prior to RIA; tritium-labeled tracer as well as iodine-labeled tracers.

Radioimmunoassay of serum parathyrin: comparison of five commercial kits.

We compared serum parathyrin radioimmunoassay data obtained with five commercially available kits for normal subjects, patients on dialysis, and patients with hyperparathyroidism. The Cambridge Nuclear kit gave low results, did not distinguish the dialysis-group sera from the normal sera, and had the greatest CV (56%) both inter- and intra-assay. The Immuno Nuclear PTH-II kit gave more satisfactory clinical data than their PTH-C-terminal kit; however, as with most procedures involving a substantial and variable nonspecific binding correction for each serum, parallelism between samples and the standard curve was poor, turnaround time slow, and inter-assay CV high (39%).

Familial occurrence of hyperuricemia, gout, and medullary cystic disease.

We observed hyperuricemia, acute gouty arthritis, and renal medullary cystic disease in three members of a family over two generations. Two of these individuals were women who developed gout by age 20 years. Two teenage sons of one of these patients had severe hyperuricemia, which appeared due to underexcretion of uric acid.

Phencyclidine-associated acute rhabdomyolysis.

Phencyclidine (PCP) is a dissociative veterinary anesthetic and tranquilizer that at present is being abused as a psychedelic and hallucinogenic agent with increasing frequency. The cases of two young patients suffering from phencyclidine toxicity are reported. In each, central nervous system depression was accompanied by an acute dystonic motor reaction resulting in acute rhabdomyolysis and myoglobinuria.