The purification and characterization of phosphonopyruvate hydrolase, a novel carbon-phosphorus bond cleavage enzyme from Variovorax sp. Pal2.

Phosphonopyruvate hydrolase, a novel bacterial carbon-phosphorus bond cleavage enzyme, was purified to homogeneity by a series of chromatographic steps from cell extracts of a newly isolated environmental strain of Variovorax sp. Pal2. The enzyme was inducible in the presence of phosphonoalanine or phosphonopyruvate; unusually, its expression was independent of the phosphate status of the cell. The native enzyme had a molecular mass of 63 kDa with a subunit mass of 31.2 kDa. Activity of purified phosphonopyruvate hydrolase was Co2+-dependent and showed a pH optimum of 6.7–7.0. The enzyme had a Km of 0.53 mM for its sole substrate, phosphonopyruvate, and was inhibited by the analogues phosphonoformic acid, 3-phosphonopropionic acid, and hydroxymethylphosphonic acid. The nucleotide sequence of the phosphonopyruvate hydrolase structural gene indicated that it is a member of the phosphoenolpyruvate phosphomutase/isocitrate lyase superfamily with 41% identity at the amino acid level to the carbon-to-phosphorus bond-forming enzyme phosphoenolpyruvate phosphomutase from Tetrahymena pyriformis. Thus its apparently ancient evolutionary origins differ from those of each of the two carbon-phosphorus hydrolases that have been reported previously; phosphonoacetaldehyde hydrolase is a member of the haloacetate dehalogenase family, whereas phosphonoacetate hydrolase belongs to the alkaline phosphatase superfamily of zinc-dependent hydrolases. Phosphonopyruvate hydrolase is likely to be of considerable significance in global phosphorus cycling, because phosphonopyruvate is known to be a key intermediate in the formation of all naturally occurring compounds that contain the carbon-phosphorus bond.

Human Papillomavirus Type 16 E6 and E7 Cause Polyploidy in Human Keratinocytes and Up-Regulation of G2-M-phase Proteins

Human papillomavirus type 16 proteins E6 and E7 have been shown to cause centrosome amplification and lagging chromosomes during mitosis. These abnormalities during mitosis can result in missegregation of the chromosomes, leading to chromosomal instability. Genomic instability is thought to be an essential part of the conversion of a normal cell to a cancer cell. We now show that E6 and E7 together cause polyploidy in primary human keratinocytes soon after these genes are introduced into the cells. Polyploidy seems to result from a spindle checkpoint failure arising from abrogation of the normal functions of p53 and retinoblastoma family members by E6 and E7, respectively. In addition, E6 and E7 cause deregulation of cellular genes such as Plk1, Aurora-A, cdk1, and Nek2, which are known to control the G2-M-phase transition and the ordered progression through mitosis.

A family of kassinatuerin-2 related peptides from the skin secretion of the African hyperoliid frog, Kassina maculata.

We describe the isolation and structural characterization of a family of antimicrobial peptides related to kassinatuerin-2, from the skin secretion of the African hyperoliid frog, Kassina maculata. All four peptides, designated kassinatuerin-2Ma through Md, are C-terminally-amidated 20-mers with the consensus sequence – FX1GAIAAALPHVIX2AIKNAL – where X1 = L/F/V/I and X2 = S/N. All four peptides are encoded by precursors of 69 amino acids. Synthetic replicates of all kassinatuerin-2 related peptides displayed a potent inhibitory activity against Staphylococcus aureus with a minimal inhibitory concentration of 16 µM, at which concentration, however, they effected 18% haemolysis of horse erythrocytes after 2 h. Despite obvious membranolytic properties, all peptides were ineffective at inhibiting the growth of Escherichia coli at concentrations up to 200 µM and were relatively ineffective against Candida albicans (MIC 120 µM). The kassinatuerin-2 related peptides of K. maculata skin secretion thus possess a discrete antimicrobial and weak haemolytic activity in contrast to the prototype kassinatuerin-2 from the skin secretion of Kassina senegalensis.

Helokinestatins: A novel family of bioactive peptides with drug-lead potential fromthe venomof the Gila Monster (Heloderma suspectum)

Venom of the Gila Monster (Heloderma suspectum) has proven to be an unlikely source of lead compounds (exendins) for the development of new injectable peptide therapeutics for the treatment of Type 2 diabetes. However, no systematic searches for new classes of bioactive peptides in lizard venom have appeared until recently. Here we describe the discovery of a new class of peptides – the helokinestatins – from H. suspectum venom, their structural characterisation and that of their biosynthetic precursors from cloned cDNA. In addition, we have subjected members of the family to preliminary pharmacological characterisation. Helokinestatins 1–6 are a family of proline-rich peptides containing 10–15 amino acid residues terminating in a common -Pro-Arg.OH motif. They are encoded in tandem within two virtually identical biosynthetic precursors of 177 and 178 amino acid residues, differing by only a single Pro residue. Each precursor also encodes a single copy of a C-type natriuretic peptide located at the C-terminus. Synthetic replicates of all helokinestatins were shown to be devoid of any direct action on the smooth muscle of rat tail artery but were found to be potent inhibitors of bradykinin-induced relaxation in this preparation in a manner that is suggestive of a non-competitive mechanism. Helokinestatin-3 (VPPPPLQMPLIPR) and helokinestatin-5 (VPPPLQMPLIPR) were found to be most potent in this respect causing almost complete inhibition of bradykinin-induced relaxation. Helokinestatins and BPPs may have a shared evolutionary history but the former do not inhibit ACE. The bradykinin inhibitory potential of helokinestatins may be exploited in the local control of chronic inflammation.

Open Adoption: Adoptive Parents' Experiences of Birth Family Contact and Talking to Their Child about Adoption

The trend towards more open adoption presents adopters with unique parenting challenges associated with satisfying the child’s curiosity about their origins, and maintaining relationships with birth family through contact. This paper focuses on the experiences of 20 sets of adoptive parents who were interviewed as part of the Northern Ireland Care Pathways and Outcomes Study. Interviews were analysed following the principles of Interpretative Phenomenological Analysis (IPA). It explores adoptive parents’ experience of talking to their child about adoption, and of post-adoption contact with members of the birth family. Adopters discussed adoption sensitively with their child, but were concerned that difficult and complex family histories would present a risk to the child’s self-esteem and emotional well-being. All forms of contact proved emotionally and practically burdensome, however adopters were committed to making it work for the child’s benefit, and were open to increased contact should the child wish it in the future. There was little relationship with birth family outside of formal contact. The study reveals a need for a mechanism to facilitate communication with birth family if adopters are to be able to respond to the child’s changing need for contact and information.

Subdivision of the bacterioferritin comigratory protein family of bacterial peroxiredoxins based on catalytic activity

Peroxiredoxins are ubiquitous proteins that catalyze the reduction of hydroperoxides, thus conferring resistance to oxidative stress. Using high-resolution mass spectrometry, we recently reclassified one such peroxiredoxin, bacterioferritin comigratory protein (BCP) of Escherichia coli, as an atypical 2-Cys peroxiredoxin that functions through the formation of an intramolecular disulfide bond between the active and resolving cysteine. An engineered E. coli BCP, which lacked the resolving cysteine, retained enzyme activity through a novel catalytic pathway. Unlike the active cysteine, the resolving cysteine of BCP peroxiredoxins is not conserved across all members of the family. To clarify the catalytic mechanism of native BCP enzymes that lack the resolving cysteine, we have investigated the BCP homologue of Burkholderia cenocepacia. We demonstrate that the B. cenocepacia BCP (BcBCP) homologue functions through a 1-Cys catalytic pathway. During catalysis, BcBCP can utilize thioredoxin as a reductant for the sulfenic acid intermediate. However, significantly higher peroxidase activity is observed utilizing glutathione as a resolving cysteine and glutaredoxin as a redox partner. Introduction of a resolving cysteine into BcBCP changes the activity from a 1-Cys pathway to an atypical 2-Cys pathway, analogous to the E. coli enzyme. In contrast to the native B. cenocepacia enzyme, thioredoxin is the preferred redox partner for this atypical 2-Cys variant. BCP-deficient B. cenocepacia exhibit a growth-phase-dependent hypersensitivity to oxidative killing. On the basis of sequence alignments, we believe that BcBCP described herein is representative of the major class of bacterial BCP peroxiredoxins. To our knowledge, this is the first detailed characterization of their catalytic activity. These studies support the subdivision of the BCP family of peroxiredoxins into two classes based on their catalytic activity.

Assessing family members' satisfaction with information sharing and communication during Hospital care at the end of life

Context: Despite the fact that most deaths occur in hospital, problems remain with how patients and families experience care at the end of life when a death occurs in a hospital. Objectives: (1) assess family member satisfaction with information sharing and communication, and (2) examine how satisfaction with information sharing and communication is associated with patient factors. Methods: Using a cross-sectional survey, data were collected from family members of adult patients who died in an acute care organization. Correlation and factor analysis were conducted, and internal consistency assessed using Cronbach's alpha. Linear regression was performed to determine the relationship among patient variables and satisfaction on the Information Sharing and Communication (ISC) scale. Results: There were 529 questionnaires available for analysis. Following correlation analysis and the dropping of redundant and conceptually irrelevant items, seven items remained for factor analysis. One factor was identified, described as information sharing and communication, that explained 76.3% of the variance. The questionnaire demonstrated good content and reliability (Cronbach's alpha 0.96). Overall, family members were satisfied with information sharing and communication (mean total satisfaction score 3.9, SD 1.1). The ISC total score was significantly associated with patient gender, the number of days in hospital before death, and the hospital program where the patient died. Conclusions: The ISC scale demonstrated good content validity and reliability. The ISC scale offers acute care organizations a means to assess the quality of information sharing and communication that transpires in care at the end of life. © Copyright 2013, Mary Ann Liebert, Inc.

First steps: study protocol for a randomized controlled trial of the effectiveness of the Group Family Nurse Partnership (gFNP) program compared to routine care in improving outcomes for high-risk mothers and their children and preventing abuse

Background: Evidence from the USA suggests that the home-based Family Nurse Partnership program (FNP), extending from early pregnancy until infants are 24 months, can reduce the risk of child abuse and neglect throughout childhood. FNP is now widely available in the UK. A new variant, Group Family Nurse Partnership (gFNP) offers similar content but in a group context and for a shorter time, until infants are 12 months old. Each group comprises 8 to 12 women with similar expected delivery dates and their partners. Its implementation has been established but there is no evidence of its effectiveness.

Methods/Design: The study comprises a multi-site randomized controlled trial designed to identify the benefits of gFNP compared to standard care. Participants (not eligible for FNP) must be either aged <20 years at their last menstrual period (LMP) with one or more previous live births, or aged 20 to 24 at LMP with low educational qualifications and no previous live births. 'Low educational qualifications' is defined as not having both Maths and English Language GCSE at grade C or higher or, if they have both, no more than four in total at grade C or higher. Exclusions are: under 20 years and previously received home-based FNP and, in either age group, severe psychotic mental illness or not able to communicate in English. Consenting women are randomly allocated (minimized by site and maternal age group) when between 10 and 16 weeks pregnant to either to the 44 session gFNP program or to standard care after the collection of baseline information. Researchers are blind to group assignment. The primary outcomes at 12 months are child abuse potential based on the revised Adult-Adolescent Parenting Inventory and parent/infant interaction coded using the CARE Index based on a video-taped interaction. Secondary outcomes are maternal depression, parenting stress, health related quality of life, social support, and use of services.

Discussion: This is the first study of the effectiveness of gFNP in the UK. Results should inform decision-making about its delivery alongside universal services, potentially enabling a wider range of families to benefit from the FNP curriculum and approach to supporting parenting.

Family-based associations in measures of psychological distress and quality of life in a cardiac screening clinic for inheritable cardiac diseases: a cross-sectional study

BACKGROUND: Family-based cardiac screening programmes for persons at risk for genetic cardiac diseases are now recommended. However, the psychological wellbeing and health related quality of life (QoL) of such screened patients is poorly understood, especially in younger patients. We sought to examine wellbeing and QoL in a representative group of adults aged 16 and over in a dedicated family cardiac screening clinic.

METHODS: Prospective survey of consecutive consenting patients attending a cardiac screening clinic, over a 12 month period. Data were collected using two health measurement tools: the Short Form 12 (version 2) and the Hospital Anxiety and Depression Scale (HADS), along with baseline demographic and screening visit-related data. The HADS and SF-12v.2 outcomes were compared by age group. Associations with a higher HADS score were examined using logistic regression, with multi-level modelling used to account for the family-based structure of the data.

RESULTS: There was a study response rate of 86.6%, with n=334 patients providing valid HADS data (valid response rate 79.5%), and data on n=316 retained for analysis. One-fifth of patients were aged under 25 (n=61). Younger patients were less likely than older to describe significant depression on their HADS scale (p<0.0001), although there were overall no difference between the prevalence of a significant HADS score between the younger and older age groups (18.0% vs 20.0%, p=0.73). Significant positive associates of a higher HADS score were having lower educational attainment, being single or separated, and being closely related to the family proband. Between-family variance in anxiety and depression scores was greater than within-family variance.

CONCLUSIONS: High levels of anxiety were seen amongst patients attending a family-based cardiac screening clinic.Younger patients also had high rates of clinically significant anxiety. Higher levels of anxiety and depression tends to run in families, and this has implications for family screening and intervention programmes.

Visible-Wavelength Survey of Jupiter-Family Cometary Nuclei as Part of SEPPCoN

We present observations of a statistically-significant number of Jupiter-family cometary nuclei as part of SEPPCoN (Survey of the Ensemble Physical Properties of Cometary Nuclei). We present preliminary results on distributions of albedos and shapes.

Homeless young people, families and change.:Family support as a facilitator to exiting homelessness.

The families of homeless young people are most often portrayed as a precipitating factor in their homelessness. However, recent studies, particularly those taking a longitudinal approach, have drawn attention to the enabling role of family members and their positive influence on the housing trajectories of homeless youth. Drawing on selected findings from an ongoing longitudinal qualitative study of homeless young people in Dublin, Ireland, this paper aims to build on this relatively fertile area of research. We demonstrate the supportive role of the families of young people who experience homelessness (often as a consequence of difficult family environments) and specifically examine how family re-engagement is negotiated and achieved. The findings highlight a number of dimensions of transition and change. Prominent among these is the importance of renewed trust and communication. Young people and their parents also had to accept responsibility for areas of life that previously served to undermine their relationships and were implicated in the circumstances surrounding a young person's premature home-leaving. Tensions and resistances on the part of young people are highlighted, demonstrating the adaptive mechanisms at work as they attempt to re-engage with family members. The implications of the findings for social work intervention with homeless youth are discussed.

Supervisor Continuity or Co-Location: Which Matters in Residency Education? Findings From a Qualitative Study of Remote Supervisor Family Physicians in Australia and Canada

Purpose: Changes to health care systems andworking hours have fragmentedresidents’ clinical experiences withpotentially negative effects ontheir development as professionals.Investigation of off-site supervision,which has been implemented in isolatedrural practice, could reveal importantbut less overt components of residencyeducation. 

Method: Insights from sociocultural learningtheory and work-based learning provideda theoretical framework. In 2011–2012,16 family physicians in Australia andCanada were asked in-depth how theyremotely supervised residents’ workand learning, and for their reflectionson this experience. The verbatiminterview transcripts and researchers’memos formed the data set. Templateanalysis produced a description andinterpretation of remote supervision. 

Results: Thirteen Australian family physiciansfrom five states and one territory, andthree Canadians from one province,participated. The main themes werehow remoteness changed the dynamicsof care and supervision; the importanceof ongoing, holistic, nonhierarchical,supportive supervisory relationships; andthat residents learned “clinical courage”through responsibility for patients’ careover time. Distance required supervisorsto articulate and pass on their expertiseto residents but made monitoringdifficult. Supervisory continuityencouraged residents to build on pastexperiences and confront deficiencies. 

Conclusions: Remote supervision enabled residents todevelop as clinicians and professionals.This questions the supremacy of co-locationas an organizing principle forresidency education. Future specialists maybenefit from programs that give themongoing and increasing responsibilityfor a group of patients and supportive.