A resilient research approach: Using community-based participatory action research in a rural area of India.

This community-based participatory action research (CBPAR) study describes a method for evaluating an after-school resilience-focused intervention in a low-resource rural area of southern India. Communities Rising, a locally developed resilience and academic program, was evaluated in a cross-continent collaboration between a research team at a U.S. university and the local community. The CBPAR literature highlights the importance of cultural considerations, community considerations, and community participation in the research process. The present case study describes the CBPAR research process and considerations at every phase of the research project, providing a road map of how community engagement can strengthen research, empower the community, and provide valuable knowledge. This study was conducted in three phases that focused on inclusion of local voices in the development both of the resilience program and the evaluation data collection process. Youth surveyors were particularly key to the research process. Data on participant demographics, satisfaction with the program, and qualitative contributions are also provided. Strengths and limitations of this study process in a rural community are discussed.

Natural products from plant cell cultures.

Plants produce complex small molecules - natural products - that exhibit anticancer, antimalarial and antimicrobial activity. These molecules play a key role in human medicine. However, plants typically produce these compounds in low quantities, and harvesting plant natural products is frequently expensive, time-consuming and environmentally damaging. Plant cell culture provides a renewable, easily scalable source of plant material. In this chapter we discuss the successes and pitfalls associated with natural product production in plant cell cultures.

Substrate specificity and diastereoselectivity of strictosidine glucosidase, a key enzyme in monoterpene indole alkaloid biosynthesis.

Strictosidine glucosidase (SGD) from Catharanthus roseus catalyzes the deglycosylation of strictosidine, an intermediate from which thousands of monoterpene indole alkaloids are derived. The steady-state kinetics of SGD with a variety of strictosidine analogs revealed the substrate preferences of this enzyme at two key positions of the strictosidine substrate. Additionally, SGD from C. roseus turns over both strictosidine and its stereoisomer vincoside, indicating that although this enzyme prefers the naturally occurring diastereomer, the enzyme is not completely diastereoselective. The implications of the substrate specificity of SGD in metabolic engineering efforts of C. roseus are highlighted.

Rapid identification of enzyme variants for reengineered alkaloid biosynthesis in periwinkle.

Monoterpene indole alkaloids from Catharanthus roseus (Madagascar periwinkle), such as the anticancer agents vinblastine and vincristine, have important pharmacological activities. Metabolic engineering of alkaloid biosynthesis can provide an efficient and environmentally friendly route to analogs of these synthetically challenging and pharmaceutically valuable natural products. However, the narrow substrate scope of strictosidine synthase, the enzyme at the entry point of the pathway, limits a pathway engineering approach. We demonstrate that with a different expression system and screening method it is possible to rapidly identify strictosidine synthase variants that accept tryptamine analogs not turned over by the wild-type enzyme. The variants are used in stereoselective synthesis of beta-carboline analogs and are assessed for biosynthetic competence within the terpene indole alkaloid pathway. These results present an opportunity to explore metabolic engineering of "unnatural" product production in the plant periwinkle.

Chemoselective derivatization of alkaloids in periwinkle.

The terpene indole alkaloid biosynthetic pathway can utilize a secologanin substrate analog containing a handle for functionalization, and the resulting non-natural alkaloids can be chemoselectively derivatized in crude extracts of plant tissue.

Directed biosynthesis of alkaloid analogs in the medicinal plant Catharanthus roseus.

Terpene indole alkaloids are plant natural products with diverse structures and biological activities. A highly branched biosynthetic pathway is responsible for the production of approximately 130 different alkaloids in Madagascar periwinkle (C. roseus) from a common biosynthetic intermediate derived from tryptamine. Although numerous biosynthetic pathways can incorporate unnatural starting materials to yield novel natural products, it was not clear how efficiently the complex, eukaryotic TIA pathway could utilize unnatural substrates to make new alkaloids. This work demonstrates that the TIA biosynthetic machinery can be used to produce novel alkaloid structures and also highlights the potential of this pathway for future metabolic engineering efforts.

Substrate specificity of strictosidine synthase.

Strictosidine synthase catalyzes a Pictet-Spengler reaction in the first step in the biosynthesis of terpene indole alkaloids to generate strictosidine. The substrate requirements for strictosidine synthase are systematically and quantitatively examined and the enzymatically generated compounds are processed by the second enzyme in this biosynthetic pathway.