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Here, new crystal structures are presented of the isolated membrane‐proximal D1 and distal D2 domains of protein tyrosine phosphatase epsilon (PTPϵ), a protein tyrosine phosphatase that has been shown to play a positive role in the survival of human breast cancer cells. A triple mutant of the PTPϵ D2 domain (A455N/V457Y/E597D) was also constructed to reconstitute the residues of the PTPϵ D1 catalytic domain that are important for phosphatase activity, resulting in only a slight increase in the phosphatase activity compared with the native D2 protein. The structures reported here are of sufficient resolution for structure‐based drug design, and a microarray‐based assay for high‐throughput screening to identify small‐molecule inhibitors of the PTPϵ D1 domain is also described. [ABSTRACT FROM AUTHOR]

The article presents information on a research which examines the effectiveness of isothiazolidinone heterocycles as inhibitors of the Yersinia pestis protein tyrosine phosphate. Also included are the materials and methods used in the research as well as a discussion on the results of the study.

The key DNA cutting and joining steps of retroviral DNA integration are carried out by the viral integrase protein. Structures of the individual domains of integrase have been determined, but their organization in the active complex with viral DNA is unknown. We show that HIV-1 integrase forms stable synaptic complexes in which a tetramer of integrase is stably associated with a pair of viral DNA ends. The viral DNA is processed within these complexes, which go on to capture the target DNA and integrate the viral DNA ends. The joining of the two viral DNA ends to target DNA occurs sequentially, with a stable intermediate complex in which only one DNA end is joined. The integration product also remains stably associated with integrase and likely requires disassembly before completion of the integration process by cellular enzymes. The results define the series of stable nucleoprotein complexes that mediate retroviral DNA integration. [ABSTRACT FROM AUTHOR]

A role for histamine in the pathogenesis of uremic pruritus was investigated in maintenance hemodialysis patients. Venous plasma histamine levels, as determined by radioenzymatic assay, were significantly higher (p < 0.05) in hemodialysis patients with pruritus (368 ± 103 pg/ml [mean ± SEM], n = 6) than in those without pruritus (146 ± 22 pg/ml, n = 5) and in normal controls (142 ± 16, n = 5). Arteriovenous fistula histamine levels (202 ± 52 pg/ml, n = 6) were significantly lower (p < 0.05) than simultaneously drawn venous samples. Markedly elevated histamine-degrading enzyme (histaminase) activities were found in both hemodialysis patients with (2.95 ± 0.18 pg histamine degraded/minute) and without (2.44 ± 0.28) pruritus, but was undetectable in normal controls. Histaminase activities did not significantly differ in simultaneously drawn venous and fistula samples. With hemodialysis, Histaminase activities fell significantly (p < 0.01), whereas plasma histamine did not change. We further examined the effects of ketotifen, a putative mast cell stabilizer, on severe uremic pruritus. Five of five patients had significant (p < 0.01) reductions in pruritus, as judged on a six-point pruritus index, after 8 weeks of drug (x = 2.3), as compared to conventional therapy (x = 5.9). Despite these improvements, no significant differences were noted in pre- versus post-drug plasma histamine levels, histaminase activities, or the histamine content per gram of skin biopsy specimen. These data support prior hypotheses that mast cell activation contributes to the pruritus of uremia. [ABSTRACT FROM AUTHOR]

We have developed competitive and direct binding methods to examine small-molecule inhibitors of protein tyrosine phosphatase activity. Focusing on the Yersinia pestis outer protein H, a potent bacterial protein tyrosine phosphatase, we describe how an understanding of the kinetic interactions involving Yersinia pestis outer protein H, peptide substrates, and small-molecule inhibitors of protein tyrosine phosphatase activity can be beneficial for inhibitor screening, and we further translate these results into a microarray assay for high-throughput screening., (Published 2012. This article is a U.S. Government work and is in the public domain in the USA.)

HIV-1 integrase (IN) is a validated therapeutic target for the treatment of AIDS. However, the emergence of resistance to raltegravir, the sole marketed FDA-approved IN inhibitor, emphasizes the need to develop second-generation inhibitors that retain efficacy against clinically relevant IN mutants. We report herein bicyclic hydroxy-1H-pyrrolopyridine-triones as a new family of HIV-1 integrase inhibitors that were efficiently prepared using a key 'Pummerer cyclization deprotonation cycloaddition' cascade of imidosulfoxides. In in vitro HIV-1 integrase assays, the analogs showed low micromolar inhibitory potencies with selectivity for strand transfer reactions as compared with 3'-processing inhibition. A representative inhibitor (5e) retained most of its inhibitory potency against the three major raltegravir-resistant IN mutant enzymes, G140S/Q148H, Y143R, and N155H. In antiviral assays employing viral vectors coding these IN mutants, compound 5e was approximately 200- and 20-fold less affected than raltegravir against the G140S/Q148H and Y143R mutations, respectively. Against the N155H mutation, 5e was approximately 10-fold less affected than raltegravir. Thus, our new compounds represent a novel structural class that may be further developed to overcome resistance to raltegravir, particularly in the case of the G140S/Q148H mutations., (© 2011 John Wiley & Sons A/S.)

Isothiazolidinone (IZD) heterocycles can act as effective components of protein tyrosine phosphatase (PTP) inhibitors by simultaneously replicating the binding interactions of both a phosphoryl group and a highly conserved water molecule, as exemplified by the structures of several PTP1B-inhibitor complexes. In the first unambiguous demonstration of IZD interactions with a PTP other than PTP1B, it is shown by X-ray crystallography that the IZD motif binds within the catalytic site of the Yersinia pestis PTP YopH by similarly displacing a highly conserved water molecule. It is also shown that IZD-based bidentate ligands can inhibit YopH in a nonpromiscuous fashion at low micromolar concentrations. Hence, the IZD moiety may represent a useful starting point for the development of YopH inhibitors.

Objective: The transcription of several cytokines, cell adhesion molecules, and enzymes involved in the inflammatory and destructive mechanisms of rheumatoid arthritis is dependent on nuclear factor kappa B (NF-kappa B). Because I kappa B kinase (IKK) is critical in transducing the signal-inducible activation of NF-kappa B, we examined whether the highly selective and orally bioavailable IKK inhibitor BMS-345541 is efficacious against collagen-induced arthritis (CIA) in mice., Methods: Arthritis in DBA/1LacJ male mice was induced by subcutaneous immunization with bovine type II collagen on day 0 and day 21. BMS-345541 was administered perorally daily, either prophylactically (before disease onset) or therapeutically (after disease onset). Clinical assessment of the incidence and severity of disease was conducted throughout the study, and histologic evaluation was performed at the time of study termination (day 42)., Results: When administered prophylactically, BMS-345541 (in a dose range of 10-100 mg/kg) was effective, in a dose-dependent manner, in reducing the incidence of disease and inhibiting clinical signs of disease. Histologic evaluation of the joints showed that both inflammation and joint destruction were blocked by the IKK inhibitor. Message levels of interleukin-1 beta in the joints were also dose-dependently inhibited in the mice that received BMS-345541. Dose-dependent efficacy in terms of both disease severity and histologic end points was observed with the therapeutic dosing regimen of BMS-345541, with use of the 100-mg/kg dose resulting in resolution of disease., Conclusion: IKK plays a key role in CIA in mice, and inhibitors of this enzyme represent a promising target for the development of novel agents to treat rheumatoid arthritis and other inflammatory diseases. BMS-345541 represents the first example of an inhibitor of IKK that has antiinflammatory activity in vivo.

Two indices of motor unit recruitment, the ramp-force and repetitive-discharge thresholds, were compared in the first dorsal interosseus muscle of ten young and twelve elderly subjects. The purpose was to determine the effect of age on the relationship between the two recruitment thresholds and the spike-triggered average force of motor units. Each subject performed three tasks requiring isometric abduction of the left index finger: a maximum voluntary contraction (MVC), a ramp-and-hold contraction, and a repetitive-discharge task. The elderly subjects used coactivation of the antagonist muscle (second palmar interosseus) more frequently than the young subjects during the ramp-and-hold contraction. Many elderly subjects expressed difficulty with the controlled ramp-down phase of the ramp-and-hold contraction and preferred a coactivation strategy to a derecruitment strategy for this task. There were no differences due to age or gender in the ramp-force thresholds between the various groups. However, the normalized repetitive-discharge threshold was significantly less for the younger subjects and for the male subjects. Nonetheless, the two recruitment thresholds were able to predict the spike-triggered average force with similar success for both the young and the elderly subjects. These data suggest that the recruitment threshold of a motor unit in first dorsal interosseus was characterized equally well by either the ramp-force or repetitive-discharge measurement for both young and elderly subjects but that coactivation was used more frequently by the elderly subjects during the ramp-and-hold task.