The role of mentoring in academic career progression:a cross-sectional survey of the Academy of Medical Sciences mentoring scheme

OBJECTIVES: To describe a successful mentoring scheme designed for mid-career clinician scientists and to examine factors associated with mentee report of positive career impact. DESIGN: Mixed methods study including in-depth interviews and cross-sectional data collection via an online survey. SETTING: Academy of Medical Sciences mentoring scheme set up in 2002 and evaluated in 2010. PARTICIPANTS: One hundred and forty-seven of 227 mentees took part in the study (response rate of 65%). Ten mentees, three mentors and eight stakeholders/scheme staff were selected to participate in in-depth interviews. MAIN OUTCOME MEASURES: Qualitative data: Interviews were transcribed, and free text was analysed to identify themes and subthemes in the narrative. Quantitative data: We examined the associations of reported positive career impact of mentoring by performing simple and multiple logistic regression analysis. RESULTS: Mentoring success was determined by a variety of factors including reasons for selection (e.g. presence of a personal recommendation), mentee characteristics (e.g. younger age), experience and skills of the mentor (e.g. ‘mentor helped me to find my own solutions’) and the quality of the relationship (e.g. ‘my mentor and I set out clear expectations early on’). CONCLUSIONS: Our evaluation demonstrates that both mentor and mentee value mentoring and that careful planning of a scheme including preparation, training and ongoing support of both mentor and mentee addressing expectations, building rapport and logistics are likely to be helpful in ensuring success and benefit from the intervention

Addition of missing loops and domains to protein models by x-ray solution scattering.

Inherent flexibility and conformational heterogeneity in proteins can often result in the absence of loops and even entire domains in structures determined by x-ray crystallographic or NMR methods. X-ray solution scattering offers the possibility of obtaining complementary information regarding the structures of these disordered protein regions. Methods are presented for adding missing loops or domains by fixing a known structure and building the unknown regions to fit the experimental scattering data obtained from the entire particle. Simulated annealing was used to minimize a scoring function containing the discrepancy between the experimental and calculated patterns and the relevant penalty terms. In low-resolution models where interface location between known and unknown parts is not available, a gas of dummy residues represents the missing domain. In high-resolution models where the interface is known, loops or domains are represented as interconnected chains (or ensembles of residues with spring forces between the C(alpha) atoms), attached to known position(s) in the available structure. Native-like folds of missing fragments can be obtained by imposing residue-specific constraints. After validation in simulated examples, the methods have been applied to add missing loops or domains to several proteins where partial structures were available

Demonstration of Proton-coupled Electron Transfer in the Copper-containing Nitrite Reductases*

The reduction of nitrite (NO2−) into nitric oxide (NO), catalyzed by nitrite reductase, is an important reaction in the denitrification pathway. In this study, the catalytic mechanism of the copper-containing nitrite reductase from Alcaligenes xylosoxidans (AxNiR) has been studied using single and multiple turnover experiments at pH 7.0 and is shown to involve two protons. A novel steady-state assay was developed, in which deoxyhemoglobin was employed as an NO scavenger. A moderate solvent kinetic isotope effect (SKIE) of 1.3 ± 0.1 indicated the involvement of one protonation to the rate-limiting catalytic step. Laser photoexcitation experiments have been used to obtain single turnover data in H2O and D2O, which report on steps kinetically linked to inter-copper electron transfer (ET). In the absence of nitrite, a normal SKIE of ∼1.33 ± 0.05 was obtained, suggesting a protonation event that is kinetically linked to ET in substrate-free AxNiR. A nitrite titration gave a normal hyperbolic behavior for the deuterated sample. However, in H2O an unusual decrease in rate was observed at low nitrite concentrations followed by a subsequent acceleration in rate at nitrite concentrations of >10 mm. As a consequence, the observed ET process was faster in D2O than in H2O above 0.1 mm nitrite, resulting in an inverted SKIE, which featured a significant dependence on the substrate concentration with a minimum value of ∼0.61 ± 0.02 between 3 and 10 mm. Our work provides the first experimental demonstration of proton-coupled electron transfer in both the resting and substrate-bound AxNiR, and two protons were found to be involved in turnover

Proton-Coupled Electron Transfer in the Catalytic Cycle of Alcaligenes xylosoxidans Copper-Dependent Nitrite Reductase

We demonstrated recently that two protons are involved in reduction of nitrite to nitric oxide through a proton-coupled electron transfer (ET) reaction catalyzed by the blue Cu-dependent nitrite reductase (Cu NiR) of Alcaligenes xylosoxidans (AxNiR). Here, the functionality of two putative proton channels, one involving Asn90 and the other His254, is studied using single (N90S, H254F) and double (N90S-H254F) mutants. All mutants studied are active, indicating that protons are still able to reach the active site. The H254F mutation has no effect on the catalytic activity, while the N90S mutation results in ∼70% decrease in activity. Laser flash-photolysis experiments show that in H254F and wild-type enzyme electrons enter at the level of the T1Cu and then redistribute between the two Cu sites. Complete ET from T1Cu to T2Cu occurs only when nitrite binds at the T2Cu site. This indicates that substrate binding to T2Cu promotes ET from T1Cu, suggesting that the enzyme operates an ordered mechanism. In fact, in the N90S and N90S-H254F variants, where the T1Cu site redox potential is elevated by ∼60 mV, inter-Cu ET is only observed in the presence of nitrite. From these results it is evident that the Asn90 channel is the main proton channel in AxNiR, though protons can still reach the active site if this channel is disrupted. Crystallographic structures provide a clear structural rationale for these observations, including restoration of the proton delivery via a significant movement of the loop connecting the T1Cu ligands Cys130 and His139 that occurs on binding of nitrite. Notably, a role for this loop in facilitating interaction of cytochrome c551 with Cu NiR has been suggested previously based on a crystal structure of the binary complex. © 2011 American Chemical Society