Group Gardening in a Native American Community: A Collaborative Approach.

Background. There is increasing awareness of the potential health benefits derived from gardening activities. Gardening practices are gaining momentum in Native American (NA) communities, yet no efforts have applied a community-based participatory research approach within a social-ecological model to understand opportunities and barriers for group gardening on an American Indian reservation. Objectives. The primary objective of this study was to identify influences across social-ecological levels that promote or hinder the implementation of community gardens and use of locally grown foods on the reservation; a secondary objective was to assess the feasibility of implementing a group gardening program for NA adults and potential of collecting health outcome measures. Method. Community members and academicians collaborated to develop and implement this study. The study (1) conducted interviews with key stakeholders to identify influences across social-ecological levels that promote or hinder the implementation of community gardens and using locally produced food and (2) assessed the physical and psychological well-being of NA adults participating in a group gardening feasibility study. Results. Major factors influencing using locally grown food and community gardens that emerged from nine interviews included knowledge/experience, self-efficacy, Elders, traditional ways, community values, generational gaps, and local tribal policies. Twenty NA adults with prediabetes or diabetes participated in the feasibility study. The Profile of Mood States Inventory showed consistently positive change in score for participants in the group gardening program versus the comparison group. Conclusions. This study identified key influences for growing locally grown food, and approaches for implementing group gardening programs for NA adults.

Parental influence on obesity in Northern Plains American Indian youth.

Little is known regarding American Indian (AI) parental influence on children's diet and physical activity (PA), or if this influence is associated with childhood weight. We compared AI parents' diet, PA, and support for these behaviors with the child's body mass index. Scores for parental support of positive PA and diet were higher among parents of overweight/obese children. Parent PA and nutrition behaviors were in a similar, but not significant, direction with respect to child body mass index. Findings suggest that future research is needed to determine what parental, societal, or community variables influence AI children to engage in healthy eating and PA, especially if they are overweight or obese, and the age at which these variables would have the most impact on these behaviors.

Amyloid formation under physiological conditions proceeds via a native-like folding intermediate.

Although most proteins can assemble into amyloid-like fibrils in vitro under extreme conditions, how proteins form amyloid fibrils in vivo remains unresolved. Identifying rare aggregation-prone species under physiologically relevant conditions and defining their structural properties is therefore an important challenge. By solving the folding mechanism of the naturally amyloidogenic protein beta-2-microglobulin at pH 7.0 and 37 degrees C and correlating the concentrations of different species with the rate of fibril elongation, we identify a specific folding intermediate, containing a non-native trans-proline isomer, as the direct precursor of fibril elongation. Structural analysis using NMR shows that this species is highly native-like but contains perturbation of the edge strands that normally protect beta-sandwich proteins from self-association. The results demonstrate that aggregation pathways can involve self-assembly of highly native-like folding intermediates, and have implications for the prevention of this, and other, amyloid disorders.

Identification of amyloidogenic peptide sequences using a coarse-grained physicochemical model.

Cross-beta amyloid is implicated in over 20 human diseases. Experiments suggest that specific sequence elements within amyloidogenic proteins play a major role in seeding amyloid formation. Identifying these seeding sequences is important for rationalizing the molecular mechanisms of amyloid formation and for elaborating therapeutic strategies that target amyloid. Theoretical techniques play an important role in facilitating the identification and structural characterization of putative seeding sequences; most amyloid species are not amenable to high resolution experimental structure techniques. In this study we have combined a coarse-grained physicochemical protein model with a highly efficient Monte Carlo sampling technique to identify amyloidogenic sequences in four proteins for which respective experimental peptide fragmentation data exist. Peptide sequences were defined as amyloidogenic if the ensemble structure predicted for three interacting peptides described a stable and regular three-stranded beta-sheet. For such peptides, free energies were calculated to provide a measure of amyloid propensity. The overall agreement between the experimental and predicted data is good, and we correctly identify several self-recognition motifs proposed to define the cross-beta amyloid fibril architectures of two of the proteins. Our results compare very favorably with those obtained using atomistic molecular dynamics methods, though our simulations are 30-40 times faster.

Time-averaged predictions of folded and misfolded peptides using a reduced physicochemical model.

Energy-based methods for calculating time-averaged peptide structures are important for rational peptide design, for defining local structure propensities in large protein chains, and for exploring the sequence determinants of amyloid formation. High-end methods are currently too slow to be practicable, and will remain so for the foreseeable future. The challenge is to create a method that runs quickly on limited computer resources and emulates reality sufficiently well. We have developed a simplified off-lattice protein model, incorporating semi-empirical physicochemical potentials, and combined it with an efficient Monte Carlo method for calculating time-averaged peptide structures. Reasonably accurate predictions are found for a set of small alpha-helical and beta-hairpin peptides, and we demonstrate a potential application in measuring local structure propensities in protein chains. Time-averaged structures have also been calculated for a set of small peptides known to form beta-amyloid fibrils. The simulations were of three interacting peptides, and in each case the time-averaged structure describes a three-stranded beta-sheet. The performance of our method in measuring the propensities of small peptides to self-associate into possible prefibrillar species compares favorably with more detailed and CPU-intensive approaches.

Water as a conformational editor in protein folding.

As molecules approach one another in aqueous solution, desolvation free energy barriers to association are encountered. Experiments suggest these (de)solvation effects contribute to the free energy barriers separating the folded and unfolded states of protein molecules. To explore their influence on the energy landscapes of protein folding reactions, we have incorporated desolvation barriers into a semi-realistic, off-lattice protein model that uses a simplified physico-chemical force-field determined solely by the sequence of amino acids. Monte Carlo sampling techniques were used to study the effects on the thermodynamics and kinetics of folding of a number of systems, diverse in structure and sequence. In each case, desolvation barriers increase the stability of the native conformation and the cooperativity of the major folding/unfolding transition. The folding times of these systems are reduced significantly upon inclusion of desolvation barriers, demonstrating that the particulate nature of the solvent engenders a more defined route to the native fold.

The disulphide mapping, folding and characterisation of recombinant Ber e 1, an allergenic protein, and SFA8, two sulphur-rich 2S plant albumins.

We have cloned and expressed genes encoding the allergenic brazil nut 2S albumin (Ber e 1) and the sunflower albumin 8 (SFA8) in the methylotrophic yeast Pichia pastoris. We show that both proteins were secreted at high levels and that the purified proteins were properly folded. We also showed that Ber e 1 is glycosylated during secretion and that the glycan does not interfere with the folding or immunoreactivity. The disulphide map of the Ber e 1 protein was experimentally established and is in agreement with the conserved disulphide structure of other members of the 2S albumin family. A model three-dimensional structure of the allergen was generated. During the expression studies and through mutation we have also shown that alteration of the sequences around the Kex2 endoproteolytic processing site in the expressed fusion protein can compromise the secretion by targeting part of the protein for possible degradation. The secreted production of these properly folded sulphur-rich plant albumins presents an opportunity to delineate the attributes that make an allergen and to facilitate the diagnosis and therapy of type I allergy.

Developing and piloting the Journey to Native Youth Health program in Northern Plains Indian communities.

PURPOSE: The purpose of this study was to develop a lifestyle change program for Native American youth by modifying the Diabetes Prevention Program (DPP) and assess implementation indicators and short term behavioral and physiological outcomes of the intervention among a small pilot sample. METHODS: Community members and project staff modified the original DPP to be developmentally and culturally appropriate for youth targeting healthy weight maintenance, lowering fat intake, and increasing physical activity. Modifications included incorporating cultural aspects and delivering the program in small groups led by community members. Sixty-four Native American youth, aged 10-14 years old were recruited from 2 Montana Indian reservations to participate in the project, titled "Journey to Native Youth Health." Participants were randomized to the Journey DPP or a health-oriented comparison condition. Pretest and posttest measures included body mass index (BMI), dietary intake, physical activity (PA), and nutrition knowledge, attitudes and beliefs (KAB). RESULTS: The target number of participants was recruited and 84% completed the program and final measures. Changes favoring the Journey DPP group were observed on measures of PA, KAB, and kilocalories from fat consumed. As expected given the short (3-month) duration of treatment, there was no overall effect on BMI at end of treatment. Among youth who were overweight or obese at baseline, however, the Journey DPP had lower BMI growth. CONCLUSIONS: Results suggest the Journey DPP is feasible to implement and has the potential to impact behaviors and weight gain associated with risk for type-2 diabetes in Native American youth.

Translating the diabetes prevention program for Northern Plains Indian youth through community-based participatory research methods.

PURPOSE: The purpose of this study was to use a community-based participatory research (CBPR) approach to translate the original Diabetes Prevention Program (DPP) to be age and culturally specific for American Indian (AI) youth. METHODS: Tribally enrolled members on 2 Montana Indian reservations conducted focus groups and interviews to discuss community members' perspectives of factors that encouraged or were barriers to healthy diet and exercise behaviors in AI youth. In total, 31 community members, aged 10 to 68 years old, participated in 4 focus groups and 14 individual interviews. Participants were self-identified as elder, cultural expert, tribal health worker, educator, parent/guardian, youth, or school food service worker. Researchers analyzed transcripts based on inductive methods of grounded theory. RESULTS: Data analysis revealed translating the DPP to youth was contingent on the lessons incorporating cultural strategies for healthy behaviors in youth such as berry picking, gardening, horseback riding, and dancing; improving knowledge and access to healthy foods and physical activity for youth and their parents; having interactive, hands-on learning activities for healthy lifestyles in the DPP lessons; using a group format and tribal members to deliver the DPP lessons; and having tribal elders talk to youth about the importance of adopting healthy behaviors when they are young. CONCLUSIONS: A CBPR approach engaged community members to identify strategies inherent in their culture, tradition, and environment that could effectively translate the DPP to Montana Indian youth living in rural reservation communities.

Trifluoromethyldiazirine: an effective photo-induced cross-linking probe for exploring amyloid formation.

The separative and analytical power of ion mobility spectrometry-mass spectrometry combined with photo-induced cross-linking of site-specifically incorporated trifluoromethyldiazirine provides a powerful approach towards structural characterisation of amyloid fibrils.

A systematic analysis of backbone amide assignments achieved via combinatorial selective labelling of amino acids.

With the advent of high-yield cell-free expressions systems, many researchers are exploiting selective isotope labelling of amino acids to increase the efficiency and accuracy of the NMR assignment process. We developed recently a combinatorial selective labelling (CSL) method capable of yielding large numbers of residue-type and sequence-specific backbone amide assignments, which involves comparing cross-peak intensities in 1H- 15N HSQC and 2D 1H- 15N HNCO spectra collected for five samples containing different combinations of 13C- and 15N-labelled amino acids [Parker MJ, Aulton-Jones M, Hounslow A, Craven C J (2004) J Am Chem Soc 126:5020-5021]. In this paper we develop a robust method for establishing the reliability of these assignments. We have performed a detailed statistical analysis of the CSL data collected for a model system (the B1 domain of protein G from Streptococcus), developing a scoring method which allows the confidence in assignments to be assessed, and which enables the effects of overlap on assignment fidelity to be predicted. To further test the scoring method and also to assess the performance of CSL in relation to sample quality, we have applied the method to the CSL data collected for GFP in our previous study.

Density guided importance sampling: application to a reduced model of protein folding.

MOTIVATION: Monte Carlo methods are the most effective means of exploring the energy landscapes of protein folding. The rugged topography of folding energy landscapes causes sampling inefficiencies however, particularly at low, physiological temperatures. RESULTS: A hybrid Monte Carlo method, termed density guided importance sampling (DGIS), is presented that overcomes these sampling inefficiencies. The method is shown to be highly accurate and efficient in determining Boltzmann weighted structural metrics of a discrete off-lattice protein model. In comparison to the Metropolis Monte Carlo method, and the hybrid Monte Carlo methods, jump-walking, smart-walking and replica-exchange, the DGIS method is shown to be more efficient, requiring no parameter optimization. The method guides the simulation towards under-sampled regions of the energy spectrum and recognizes when equilibrium has been reached, avoiding arbitrary and excessively long simulation times. AVAILABILITY: Fortran code available from authors upon request. CONTACT: m.j.parker@leeds.ac.uk.

A combinatorial selective labeling method for the assignment of backbone amide NMR resonances.

A combinatorial selective labeling (CSL) method is presented for the assignment of backbone amide NMR resonances, which has a particular application in the identification of protein-ligand interaction sites. The method builds on the dual amino acid selective labeling technique. In the CSL method a number of different samples are produced, each with a different pattern of labeled amino acids. By analyzing peak intensities in HSQC and 2D HNCO spectra of these samples, a large number of combinations of amino acid pairs can be simultaneously assigned. We demonstrate the method on the 27 kDa protein GFP. The samples can be produced rapidly and cost-effectively in a commercially available in vitro translation system. The method greatly simplifies the process of backbone assignment and would be very straightforward to automate.