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1.
Appl Microbiol Biotechnol ; 104(13): 6013-6022, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32367311

RESUMO

Directed evolution has become an important method to unleash the latent potential of enzymes to make them uniquely suited for human purposes. However, the need for a large reagent volume and sophisticated instrumentation hampers its broad implementation. In an attempt to address this problem, here we report a paper-based high-throughput screening approach that should find broad application in generating desired enzymes. As an example case, the dehalogenation reaction of the halohydrin dehalogenase was adopted for assay development. In addition to visual detection, quantitative measurements were performed by measuring the color intensity of an image that was photographed by a smartphone and processed using ImageJ free software. The proposed method was first validated using a gold standard method and then applied to mutagenesis library screening with reduced consumption of reagents (i.e., ≤ 10 µl per assay) and a shorter assay time. We identified two active mutants (P135A and G137A) with improved activities toward four tested substrates. The assay not only consumes less reagents but also eliminates the need for expensive instrumentation. The proposed method demonstrates the potential of paper-based whole-cell screening coupled with digital image colorimetry as a promising approach for the discovery of industrially important enzymes.Key Points• A frugal method was developed for directed enzyme evolution.• Mutagenesis libraries were successfully screened on a paper platform.• Smartphone imaging was efficiently used to measure enzyme activities.


Assuntos
Evolução Molecular Direcionada/métodos , Ensaios de Triagem em Larga Escala/métodos , Papel , Catálise , Colorimetria , Evolução Molecular Direcionada/economia , Evolução Molecular Direcionada/normas , Escherichia coli/genética , Escherichia coli/metabolismo , Biblioteca Gênica , Ensaios de Triagem em Larga Escala/economia , Ensaios de Triagem em Larga Escala/normas , Hidrolases/genética , Hidrolases/metabolismo , Mutagênese , Mutação , Reprodutibilidade dos Testes , Smartphone
2.
Curr Dev Nutr ; 2(12): nzy063, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30569029

RESUMO

Although significant achievements in human health have been made globally, progress has been made possible, in part, through unconstrained use of natural resources. As the health of our planet worsens, human health is also endangered. Scholars and policymakers from diverse disciplines highlighted complex, multisectoral approaches for addressing poor dietary intake, over- and undernutrition, and chronic diseases in sub-Saharan Africa at the Agriculture, Nutrition, Health, and the Environment in Africa Conference held at Harvard University on 6-7 November 2017. A planetary health approach to addressing these challenges offers a unique opportunity to advance solutions for environmental and social factors that influence agriculture, nutrition, and overall health in the larger context of rapid population growth and transitions in food systems and livelihoods. This paper outlines 3 key avenues for universities to promote science at the intersection of public health and the environment in sub-Saharan Africa.

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