Iterative approach to computational enzyme design

A general approach for the computational design of enzymes to catalyze arbitrary reactions is a goal at the forefront of the field of protein design. Recently, computationally designed enzymes have been produced for three chemical reactions through the synthesis and screening of a large number of variants. Here, we present an iterative approach that has led to the development of the most catalytically efficient computationally designed enzyme for the Kemp elimination to date. Previously established computational techniques were used to generate an initial design, HG-1, which was catalytically inactive. Analysis of HG-1 with molecular dynamics simulations (MD) and X-ray crystallography indicated that the inactivity might be due to bound waters and high flexibility of residues within the active site. This analysis guided changes to our design procedure, moved the design deeper into the interior of the protein, and resulted in an active Kemp eliminase, HG-2. The cocrystal structure of this enzyme with a transition state analog (TSA) revealed that the TSA was bound in the active site, interacted with the intended catalytic base in a catalytically relevant manner, but was flipped relative to the design model. MD analysis of HG-2 led to an additional point mutation, HG-3, that produced a further threefold improvement in activity. This iterative approach to computational enzyme design, including detailed MD and structural analysis of both active and inactive designs, promises a more complete understanding of the underlying principles of enzymatic catalysis and furthers progress toward reliably producing active enzymes

Services for reducing duration of hospital care for acute stroke patients

<p>Background: Stroke patients conventionally receive a substantial part of their rehabilitation in hospital. Services have now been developed which offer patients in hospital an early discharge with rehabilitation at home (early supported discharge (ESD)).</p> <p>Objectives: To establish the effects and costs of ESD services compared with conventional services.</p> <p>Search methods: We searched the trials registers of the Cochrane Stroke Group (January 2012) and the Cochrane Effective Practice and Organisation of Care (EPOC) Group, MEDLINE (2008 to 7 February 2012), EMBASE (2008 to 7 February 2012) and CINAHL (1982 to 7 February 2012). In an effort to identify further published, unpublished and ongoing trials we searched 17 trial registers (February 2012), performed citation tracking of included studies, checked reference lists of relevant articles and contacted trialists.</p> <p>Selection criteria: Randomised controlled trials recruiting stroke patients in hospital to receive either conventional care or any service intervention which has provided rehabilitation and support in a community setting with an aim of reducing the duration of hospital care.</p> <p>Data collection and analysis: The primary patient outcome was the composite end-point of death or long-term dependency recorded at the end of scheduled follow up. Two review authors scrutinised trials and categorised them on their eligibility.We then sought standardised individual patient data from the primary trialists. We analysed the results for all trials and for subgroups of patients and services, in particular whether the intervention was provided by a co-ordinated multidisciplinary team (co-ordinated ESD team) or not.</p> <p>Main results: Outcome data are currently available for 14 trials (1957 patients). Patients tended to be a selected elderly group withmoderate disability. The ESD group showed significant reductions (P = 0.0001) in the length of hospital stay equivalent to approximately seven days. Overall, the odds ratios (OR) (95% confidence interval (CI)) for death, death or institutionalisation, death or dependency at the end of scheduled follow-up were OR 0.91 (95% CI 0.67 to 1.25, P = 0.58), OR 0.78 (95% CI 0.61 to 1.00, P = 0.05) and OR 0.80 (95% CI 0.67 to 0.97, P = 0.02) respectively. The greatest benefits were seen in the trials evaluating a co-ordinated ESD team and in stroke patients with mild to moderate disability. Improvements were also seen in patients’ extended activities of daily living scores (standardised mean difference 0.12, 95% CI 0.00 to 0.25, P = 0.05) and satisfaction with services (OR 1.60, 95% CI 1.08 to 2.38, P = 0.02) but no statistically significant differences were seen in carers’ subjective health status, mood or satisfaction with services. The apparent benefits were no longer statistically significant at five-year follow-up.</p> <p>Authors’ conclusions: Appropriately resourced ESD services provided for a selected group of stroke patients can reduce long-term dependency and admission to institutional care as well as reducing the length of hospital stay. We observed no adverse impact on the mood or subjective health status of patients or carers.</p&gt

Con A induced suppressor cells: Suppression with I region incompatibility

Allogeneic Con A induced suppressor cells differing in the I and S region but not H-2K or H-2D regions were as efficient as syngeneic cells in suppressing the secondary IgM and IgG response to burro erythrocytes. Con A activated suppressor cells were not sensitive to anti-Ia serum and complement. However, if the spleen cell population was treated with anti-Ia serum and complement before stimulation with Con A, suppressor cells were not generated.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/22960/1/0000527.pd

The Freshman, vol. 5, no. 17

The Freshman was a weekly, student newsletter issued on Mondays throughout the academic year. The newsletter included calendar notices, coverage of campus social events, lectures, and athletic teams. The intent of the publication was to create unity, a sense of community, and class spirit among first year students

Nonsusceptibility of Primate Cells to Taura Syndrome Virus

Primate cells commonly used to test for viruses of the Picornaviridae family are not susceptible to infection by Taura syndrome virus of penaeid shrimp

Pan-cancer Alterations of the MYC Oncogene and Its Proximal Network across the Cancer Genome Atlas

Although theMYConcogene has been implicated incancer, a systematic assessment of alterations ofMYC, related transcription factors, and co-regulatoryproteins, forming the proximal MYC network (PMN),across human cancers is lacking. Using computa-tional approaches, we define genomic and proteo-mic features associated with MYC and the PMNacross the 33 cancers of The Cancer Genome Atlas.Pan-cancer, 28% of all samples had at least one ofthe MYC paralogs amplified. In contrast, the MYCantagonists MGA and MNT were the most frequentlymutated or deleted members, proposing a roleas tumor suppressors.MYCalterations were mutu-ally exclusive withPIK3CA,PTEN,APC,orBRAFalterations, suggesting that MYC is a distinct onco-genic driver. Expression analysis revealed MYC-associated pathways in tumor subtypes, such asimmune response and growth factor signaling; chro-matin, translation, and DNA replication/repair wereconserved pan-cancer. This analysis reveals insightsinto MYC biology and is a reference for biomarkersand therapeutics for cancers with alterations ofMYC or the PMN

A multidisciplinary, multifactorial intervention program reduces postoperative falls and injuries after femoral neck fracture

INTRODUCTION: This study evaluates whether a postoperative multidisciplinary, intervention program, including systematic assessment and treatment of fall risk factors, active prevention, detection, and treatment of postoperative complications, could reduce inpatient falls and fall-related injuries after a femoral neck fracture. METHODS: A randomized, controlled trial at the orthopedic and geriatric departments at Umeå University Hospital, Sweden, included 199 patients with femoral neck fracture, aged  ≥70 years. RESULTS: Twelve patients fell 18 times in the intervention group compared with 26 patients suffering 60 falls in the control group. Only one patient with dementia fell in the intervention group compared with 11 in the control group. The crude postoperative fall incidence rate was 6.29/1,000 days in the intervention group vs 16.28/1,000 days in the control group. The incidence rate ratio was 0.38 [95% confidence interval (CI): 0.20 – 0.76, p = 0.006] for the total sample and 0.07 (95% CI: 0.01–0.57, p=0.013) among patients with dementia. There were no new fractures in the intervention group but four in the control group. CONCLUSION: A team applying comprehensive geriatric assessment and rehabilitation, including prevention, detection, and treatment of fall risk factors, can successfully prevent inpatient falls and injuries, even in patients with dementia

Bioterrorism-related Inhalational Anthrax in an Elderly Woman, Connecticut, 2001

On November 20, 2001, inhalational anthrax was confirmed in an elderly woman from rural Connecticut. To determine her exposure source, we conducted an extensive epidemiologic, environmental, and laboratory investigation. Molecular subtyping showed that her isolate was indistinguishable from isolates associated with intentionally contaminated letters. No samples from her home or community yielded Bacillus anthracis, and she received no first-class letters from facilities known to have processed intentionally contaminated letters. Environmental sampling in the regional Connecticut postal facility yielded B. anthracis spores from 4 (31%) of 13 sorting machines. One extensively contaminated machine primarily processes bulk mail. A second machine that does final sorting of bulk mail for her zip code yielded B. anthracis on the column of bins for her carrier route. The evidence suggests she was exposed through a cross-contaminated bulk mail letter. Such cross-contamination of letters and postal facilities has implications for managing the response to future B. anthracis–contaminated mailings

Heterozygosity for Pten Promotes Tumorigenesis in a Mouse Model of Medulloblastoma

BACKGROUND: Recent publications have described an important role for cross talk between PI-3 kinase and sonic hedgehog signaling pathways in the pathogenesis of medulloblastoma. METHODOLOGY/PRINCIPAL FINDINGS: We crossed mice with constitutive activation of Smoothened, SmoA1, with Pten deficient mice. Both constitutive and conditional Pten deficiency doubled the incidence of mice with symptoms of medulloblastoma and resulted in decreased survival. Analysis revealed a clear separation of gene signatures, with up-regulation of genes in the PI-3 kinase signaling pathway, including downstream activation of angiogenesis in SmoA1+/-; Pten +/- medulloblastomas. Western blotting and immunohistochemistry confirmed reduced or absent Pten, Akt activation, and increased angiogenesis in Pten deficient tumors. Down-regulated genes included genes in the sonic hedgehog pathway and tumor suppressor genes. SmoA1+/-; Pten +/+ medulloblastomas appeared classic in histology with increased proliferation and diffuse staining for apoptosis. In contrast, Pten deficient tumors exhibited extensive nodularity with neuronal differentiation separated by focal areas of intense staining for proliferation and virtually absent apoptosis. Examination of human medulloblastomas revealed low to absent PTEN expression in over half of the tumors. Kaplan-Meier analysis confirmed worse overall survival in patients whose tumor exhibited low to absent PTEN expression. CONCLUSIONS/SIGNIFICANCE: This suggests that PTEN expression is a marker of favorable prognosis and mouse models with activation of PI-3 kinase pathways may be important tools for preclinical evaluation of promising agents for the treatment of medulloblastoma