ASU Scholarship Showcase

Photosynthesis, a process catalysed by plants, algae and cyanobacteria converts sunlight to energy thus sustaining all higher life on Earth. Two large membrane protein complexes, photosystem I and II (PSI and PSII), act in series to catalyse the light-driven reactions in photosynthesis. PSII catalyses the light-driven water splitting process, which maintains the Earth’s oxygenic atmosphere. In this process, the oxygen-evolving complex (OEC) of PSII cycles through five states, S₀ to S₄, in which four electrons are sequentially extracted from the OEC in four light-driven charge-separation events. Here we describe time resolved experiments on PSII nano/microcrystals from Thermosynechococcus elongatus performed with the recently developed technique of serial femtosecond crystallography. Structures have been determined from PSII in the dark S₁ state and after double laser excitation (putative S₃ state) at 5 and 5.5 Å resolution, respectively. The results provide evidence that PSII undergoes significant conformational changes at the electron acceptor side and at the Mn₄CaO₅ core of the OEC. These include an elongation of the metal cluster, accompanied by changes in the protein environment, which could allow for binding of the second substrate water molecule between the more distant protruding Mn (referred to as the ‘dangler’ Mn) and the Mn₃CaO_x cubane in the S₂ to S₃ transition, as predicted by spectroscopic and computational studies. This work shows the great potential for time-resolved serial femtosecond crystallography for investigation of catalytic processes in biomolecules.

We present results from experiments at the Linac Coherent Light Source (LCLS) demonstrating that serial femtosecond crystallography (SFX) can be performed to high resolution (~2.5 Å) using protein microcrystals deposited on an ultra-thin silicon nitride membrane and embedded in a preservation medium at room temperature. Data can be acquired at a high acquisition rate using x-ray free electron laser sources to overcome radiation damage, while sample consumption is dramatically reduced compared to flowing jet methods. We achieved a peak data acquisition rate of 10 Hz with a hit rate of ~38%, indicating that a complete data set could be acquired in about one 12-hour LCLS shift using the setup described here, or in even less time using hardware optimized for fixed target SFX. This demonstration opens the door to ultra low sample consumption SFX using the technique of diffraction-before-destruction on proteins that exist in only small quantities and/or do not produce the copious quantities of microcrystals required for flowing jet methods.

Background: Worksites are important locations for interventions to promote health. However, occupational programs with documented efficacy often are not used, and those being implemented have not been studied. The research in this report was funded through the American Reinvestment and Recovery Act Challenge Topic 'Pathways for Translational Research,' to define and prioritize determinants that enable and hinder translation of evidenced-based health interventions in well-defined settings.

Methods: The IGNITE (investigation to guide new insights for translational effectiveness) trial is a prospective cohort study of a worksite wellness and injury reduction program from adoption to final outcomes among 12 fire departments. It will employ a mixed methods strategy to define a translational model. We will assess decision to adopt, installation, use, and outcomes (reach, individual outcomes, and economic effects) using onsite measurements, surveys, focus groups, and key informant interviews. Quantitative data will be used to define the model and conduct mediation analysis of each translational phase. Qualitative data will expand on, challenge, and confirm survey findings and allow a more thorough understanding and convergent validity by overcoming biases in qualitative and quantitative methods used alone.

Discussion: Findings will inform worksite wellness in fire departments. The resultant prioritized influences and model of effective translation can be validated and manipulated in these and other settings to more efficiently move science to service.

The number and variety of connectivity estimation methods is likely to continue to grow over the coming decade. Comparisons between methods are necessary to prune this growth to only the most accurate and robust methods. However, the nature of connectivity is elusive with different methods potentially attempting to identify different aspects of connectivity. Commonalities of connectivity definitions across methods upon which base direct comparisons can be difficult to derive. Here, we explicitly define “effective connectivity” using a common set of observation and state equations that are appropriate for three connectivity methods: dynamic causal modeling (DCM), multivariate autoregressive modeling (MAR), and switching linear dynamic systems for fMRI (sLDSf). In addition while deriving this set, we show how many other popular functional and effective connectivity methods are actually simplifications of these equations. We discuss implications of these connections for the practice of using one method to simulate data for another method. After mathematically connecting the three effective connectivity methods, simulated fMRI data with varying numbers of regions and task conditions is generated from the common equation. This simulated data explicitly contains the type of the connectivity that the three models were intended to identify. Each method is applied to the simulated data sets and the accuracy of parameter identification is analyzed. All methods perform above chance levels at identifying correct connectivity parameters. The sLDSf method was superior in parameter estimation accuracy to both DCM and MAR for all types of comparisons.

Previous proof-of-concept measurements on single-layer two-dimensional membrane-protein crystals performed at X-ray free-electron lasers (FELs) have demonstrated that the collection of meaningful diffraction patterns, which is not possible at synchrotrons because of radiation-damage issues, is feasible. Here, the results obtained from the analysis of a thousand single-shot, room-temperature X-ray FEL diffraction images from two-dimensional crystals of a bacteriorhodopsin mutant are reported in detail. The high redundancy in the measurements boosts the intensity signal-to-noise ratio, so that the values of the diffracted intensities can be reliably determined down to the detector-edge resolution of 4 Å. The results show that two-dimensional serial crystallography at X-ray FELs is a suitable method to study membrane proteins to near-atomic length scales at ambient temperature. The method presented here can be extended to pump–probe studies of optically triggered structural changes on submillisecond timescales in two-dimensional crystals, which allow functionally relevant large-scale motions that may be quenched in three-dimensional crystals.

Background: Counselor behaviors that mediate the efficacy of motivational interviewing (MI) are not well understood, especially when applied to health behavior promotion. We hypothesized that client change talk mediates the relationship between counselor variables and subsequent client behavior change.

Methods: Purposeful sampling identified individuals from a prospective randomized worksite trial using an MI intervention to promote firefighters’ healthy diet and regular exercise that increased dietary intake of fruits and vegetables (n = 21) or did not increase intake of fruits and vegetables (n = 22). MI interactions were coded using the Motivational Interviewing Skill Code (MISC 2.1) to categorize counselor and firefighter verbal utterances. Both Bayesian and frequentist mediation analyses were used to investigate whether client change talk mediated the relationship between counselor skills and behavior change.

Results: Counselors’ global spirit, empathy, and direction and MI-consistent behavioral counts (e.g., reflections, open questions, affirmations, emphasize control) significantly correlated with firefighters’ total client change talk utterances (rs = 0.42, 0.40, 0.30, and 0.61, respectively), which correlated significantly with their fruit and vegetable intake increase (r = 0.33). Both Bayesian and frequentist mediation analyses demonstrated that findings were consistent with hypotheses, such that total client change talk mediated the relationship between counselor’s skills—MI-consistent behaviors [Bayesian mediated effect: αβ = .06 (.03), 95% CI = .02, .12] and MI spirit [Bayesian mediated effect: αβ = .06 (.03), 95% CI = .01, .13]—and increased fruit and vegetable consumption.

Conclusion: Motivational interviewing is a resource- and time-intensive intervention, and is currently being applied in many arenas. Previous research has identified the importance of counselor behaviors and client change talk in the treatment of substance use disorders. Our results indicate that similar mechanisms may underlie the effects of MI for dietary change. These results inform MI training and application by identifying those processes critical for MI success in health promotion domains.

Background: Carriers of the APOE ε4 allele are at increased risk of developing Alzheimer’s disease (AD), and have been shown to have reduced cerebral metabolic rate of glucose (CMRgl) in the same brain areas frequently affected in AD. These individuals also exhibit reduced plasma levels of apolipoprotein E (apoE) attributed to a specific decrease in the apoE4 isoform as determined by quantification of individual apoE isoforms in APOE ε4 heterozygotes. Whether low plasma apoE levels are associated with structural and functional brain measurements and cognitive performance remains to be investigated.

Methods: Using quantitative mass spectrometry we quantified the plasma levels of total apoE and the individual apoE3 and apoE4 isoforms in 128 cognitively normal APOE ε3/ε4 individuals included in the Arizona APOE cohort. All included individuals had undergone extensive neuropsychological testing and 25 had in addition undergone FDG-PET and MRI to determine CMRgl and regional gray matter volume (GMV).

Results: Our results demonstrated higher apoE4 levels in females versus males and an age-dependent increase in the apoE3 isoform levels in females only. Importantly, a higher relative ratio of apoE4 over apoE3 was associated with GMV loss in the right posterior cingulate and with reduced CMRgl bilaterally in the anterior cingulate and in the right hippocampal area. Additional exploratory analysis revealed several negative associations between total plasma apoE, individual apoE isoform levels, GMV and CMRgl predominantly in the frontal, occipital and temporal areas. Finally, our results indicated only weak associations between apoE plasma levels and cognitive performance which further appear to be affected by sex.

Conclusions: Our study proposes a sex-dependent and age-dependent variation in plasma apoE isoform levels and concludes that peripheral apoE levels are associated with GMV, CMRgl and possibly cognitive performance in cognitively healthy individuals with a genetic predisposition to AD.

It has been suggested that the extended intensity profiles surrounding Bragg reflections that arise when a series of finite crystals of varying size and shape are illuminated by the intense, coherent illumination of an x-ray free-electron laser may enable the crystal’s unit-cell electron density to be obtained ab initio via well-established iterative phasing algorithms. Such a technique could have a significant impact on the field of biological structure determination since it avoids the need for a priori information from similar known structures, multiple measurements near resonant atomic absorption energies, isomorphic derivative crystals, or atomic-resolution data. Here, we demonstrate this phasing technique on diffraction patterns recorded from artificial two-dimensional microcrystals using the seeded soft x-ray free-electron laser FERMI. We show that the technique is effective when the illuminating wavefront has nonuniform phase and amplitude, and when the diffraction intensities cannot be measured uniformly throughout reciprocal space because of a limited signal-to-noise ratio.

The objective of this study was to identify physical, social, and intrapersonal cues that were associated with the consumption of sweetened beverages and sweet and salty snacks among adolescents from lower SES neighborhoods. Students were recruited from high schools with a minimum level of 25% free or reduced cost lunches. Using ecological momentary assessment, participants (N = 158) were trained to answer brief questionnaires on handheld PDA devices: (a) each time they ate or drank, (b) when prompted randomly, and (c) once each evening. Data were collected over 7 days for each participant. Participants reported their location (e.g., school grounds, home), mood, social environment, activities (e.g., watching TV, texting), cravings, food cues (e.g., saw a snack), and food choices. Results showed that having unhealthy snacks or sweet drinks among adolescents was associated with being at school, being with friends, feeling lonely or bored, craving a drink or snack, and being exposed to food cues. Surprisingly, sweet drink consumption was associated with exercising. Watching TV was associated with consuming sweet snacks but not with salty snacks or sweet drinks. These findings identify important environmental and intrapersonal cues to poor snacking choices that may be applied to interventions designed to disrupt these food-related, cue-behavior linked habits.

Methodologists have developed mediation analysis techniques for a broad range of substantive applications, yet methods for estimating mediating mechanisms with missing data have been understudied. This study outlined a general Bayesian missing data handling approach that can accommodate mediation analyses with any number of manifest variables. Computer simulation studies showed that the Bayesian approach produced frequentist coverage rates and power estimates that were comparable to those of maximum likelihood with the bias-corrected bootstrap. We share an SAS macro that implements Bayesian estimation and use 2 data analysis examples to demonstrate its use.