Implementation evaluation of a collective impact initiative to promote adolescent health in Oklahoma County, USA.

BACKGROUND: The teenage birth rate in the USA has considerably decreased in recent decades; however, more innovative, collaborative approaches are needed to promote adolescent health and prevent teenage pregnancy at the community level. Despite literature on the promising results of the collective impact (CI) model for health promotion, there is limited literature on the model's ability to reduce teenage pregnancies in a community. The Central Oklahoma Teen Pregnancy Prevention Collaboration is applying the CI model to foster collaboration among multiple stakeholders with the goal of increasing community and organizational capacity to improve adolescent health outcomes. This paper reports the findings from the initiative's implementation evaluation, which sought to understand whether the CI model improved collaboration among organizations and understand barriers and facilitators that affected program delivery. METHODS: Program implementers and evaluators jointly developed research questions to guide the intervention and evaluation design. The Consolidated Framework for Implementation Research (CFIR) was used to assess program components including the intervention characteristics, organization setting, community setting, facilitator characteristics, and the process of implementation. Primary sources of data included performance measures, meeting observations (n = 11), and semi-structured interviews (n = 10). The data was thematically analyzed using CFIR constructs, community capacity domains, and the five constructs of CI. RESULTS: Key findings include the need for shortened meeting times for meaningful engagement, opportunities for organizations to take on more active roles in the Collaboration, and enhanced community context expertise (i.e., those with lived experience) in all Collaboration initiatives. We identified additional elements to the core constructs of CI that are necessary for successful implementation: distinct role identification for partner organizations and incorporation of equity and inclusivity into collaboration processes and procedures. CONCLUSIONS: Results from this implementation evaluation provide valuable insights into implementation fidelity, participant experience, and implementation reach of an innovative, systems-level program. Findings demonstrate the context and requirements needed to successfully implement this innovative program approach and CI overall. Additional core elements for CI are identified and contribute to the growing body of literature on successful CI initiatives.

Creation of a formate: malate oxidoreductase by fusion of dehydrogenase enzymes with PEGylated cofactor swing arms.

Enzymatic biocatalysis can be limited by the necessity of soluble cofactors. Here, we introduced PEGylated nicotinamide adenine dinucleotide (NAD(H)) swing arms to two covalently fused dehydrogenase enzymes to eliminate their nicotinamide cofactor requirements. A formate dehydrogenase and cytosolic malate dehydrogenase were connected via SpyCatcher-SpyTag fusions. Bifunctionalized polyethylene glycol chains tethered NAD(H) to the fusion protein. This produced a formate:malate oxidoreductase that exhibited cofactor-independent ping-pong kinetics with predictable Michaelis constants. Kinetic modeling was used to explore the effective cofactor concentrations available for electron transfer in the complexes. This approach could be used to create additional cofactor-independent transhydrogenase biocatalysts by swapping fused dehydrogenases.

Direct Evidence for Metabolon Formation and Substrate Channeling in Recombinant TCA Cycle Enzymes.

Supramolecular assembly of enzymes into metabolon structures is thought to enable efficient transport of reactants between active sites via substrate channeling. Recombinant versions of porcine citrate synthase (CS), mitochondrial malate dehydrogenase (mMDH), and aconitase (Aco) were found to adopt a homogeneous native-like metabolon structure in vitro. Site-directed mutagenesis performed on highly conserved arginine residues located in the positively charged channel connecting mMDH and CS active sites led to the identification of CS(R65A) which retained high catalytic efficiency. Substrate channeling between the CS mutant and mMDH was severely impaired and the overall channeling probability decreased from 0.99 to 0.023. This work provides direct mechanistic evidence for the channeling of reaction intermediates, and disruption of this interaction would have important implications on the control of flux in central carbon metabolism.

Designed protein aggregates entrapping carbon nanotubes for bioelectrochemical oxygen reduction.

The engineering of robust protein/nanomaterial interfaces is critical in the development of bioelectrocatalytic systems. We have used computational protein design to identify two amino acid mutations in the small laccase protein (SLAC) from Streptomyces coelicolor to introduce new inter-protein disulfide bonds. The new dimeric interface introduced by these disulfide bonds in combination with the natural trimeric structure drive the self-assembly of SLAC into functional aggregates. The mutations had a minimal effect on kinetic parameters, and the enzymatic assemblies exhibited an increased resistance to irreversible thermal denaturation. The SLAC assemblies were combined with single-walled carbon nanotubes (SWNTs), and explored for use in oxygen reduction electrodes. The incorporation of SWNTs into the SLAC aggregates enabled operation at an elevated temperature and reduced the reaction overpotential. A current density of 1.1 mA/cm(2) at 0 V versus Ag/AgCl was achieved in an air-breathing cathode system. Biotechnol. Bioeng. 2016;113: 2321-2327. © 2016 Wiley Periodicals, Inc.

Functional interfaces for biomimetic energy harvesting: CNTs-DNA matrix for enzyme assembly.

The development of 3D structures exploring the properties of nano-materials and biological molecules has been shown through the years as an effective path forward for the design of advanced bio-nano architectures for enzymatic fuel cells, photo-bio energy harvesting devices, nano-biosensors and bio-actuators and other bio-nano-interfacial architectures. In this study we demonstrate a scaffold design utilizing carbon nanotubes, deoxyribose nucleic acid (DNA) and a specific DNA binding transcription factor that allows for directed immobilization of a single enzyme. Functionalized carbon nanotubes were covalently bonded to a diazonium salt modified gold surface through carbodiimide chemistry creating a brush-type nanotube alignment. The aligned nanotubes created a highly ordered structure with high surface area that allowed for the attachment of a protein assembly through a designed DNA scaffold. The enzyme immobilization was controlled by a zinc finger (ZNF) protein domain that binds to a specific dsDNA sequence. ZNF 268 was genetically fused to the small laccase (SLAC) from Streptomyces coelicolor, an enzyme belonging to the family of multi-copper oxidases, and used to demonstrate the applicability of the developed approach. Analytical techniques such as X-ray photoelectron spectroscopy (XPS), atomic force microscopy (AFM), and enzymatic activity analysis, allowed characterization at each stage of development of the bio-nano architecture. This article is part of a Special Issue entitled Biodesign for Bioenergetics--the design and engineering of electronic transfer cofactors, proteins and protein networks, edited by Ronald L. Koder and J.L. Ross Anderson.

Enzymatic fuel cells: integrating flow-through anode and air-breathing cathode into a membrane-less biofuel cell design.

One of the key goals of enzymatic biofuel cells research has been the development of a fully enzymatic biofuel cell that operates under a continuous flow-through regime. Here, we present our work on achieving this task. Two NAD(+)-dependent dehydrogenase enzymes; malate dehydrogenase (MDH) and alcohol dehydrogenase (ADH) were independently coupled with poly-methylene green (poly-MG) catalyst for biofuel cell anode fabrication. A fungal laccase that catalyzes oxygen reduction via direct electron transfer (DET) was used as an air-breathing cathode. This completes a fully enzymatic biofuel cell that operates in a flow-through mode of fuel supply polarized against an air-breathing bio-cathode. The combined, enzymatic, MDH-laccase biofuel cell operated with an open circuit voltage (OCV) of 0.584 V, whereas the ADH-laccase biofuel cell sustained an OCV of 0.618 V. Maximum volumetric power densities approaching 20 µW cm(-3) are reported, and characterization criteria that will aid in future optimization are discussed.