Structural basis for native agonist and synthetic inhibitor recognition by the Pseudomonas aeruginosa quorum sensing regulator PqsR (MvfR)

Bacterial populations co-ordinate gene expression collectively through quorum sensing (QS), a cell-to-cell communication mechanism employing diffusible signal molecules. The LysR-type transcriptional regulator (LTTR) protein PqsR (MvfR) is a key component of alkyl-quinolone (AQ)-dependent QS in Pseudomonas aeruginosa. PqsR is activated by 2-alkyl-4-quinolones including the Pseudomonas quinolone signal (PQS; 2-heptyl-3-hydroxy-4(1H)-quinolone), its precursor 2-heptyl-4- hydroxyquinoline (HHQ) and their C9 congeners, 2-nonyl-3-hydroxy-4(1H)-quinolone (C9-PQS) and 2-nonyl-4-hydroxyquinoline (NHQ). These drive the autoinduction of AQ biosynthesis and the up-regulation of key virulence determinants as a function of bacterial population density. Consequently, PqsR constitutes a potential target for novel antibacterial agents which attenuate infection through the blockade of virulence. Here we present the crystal structures of the PqsR co-inducer binding domain (CBD) and a complex with the native agonist NHQ. We show that the structure of the PqsR CBD has an unusually large ligand-binding pocket in which a native AQ agonist is stabilized entirely by hydrophobic interactions. Through a ligand-based design strategy we synthesized and evaluated a series of 50 AQ and novel quinazolinone (QZN) analogues and measured the impact on AQ biosynthesis, virulence gene expression and biofilm development. The simple exchange of two isosteres (OH for NH2) switches a QZN agonist to an antagonist with a concomitant impact on the induction of bacterial virulence factor production. We also determined the complex crystal structure of a QZN antagonist bound to PqsR revealing a similar orientation in the ligand binding pocket to the native agonist NHQ. This structure represents the first description of an LTTR-antagonist complex. Overall these studies present novel insights into LTTR ligand binding and ligand-based drug design and provide a chemical scaffold for further anti-P. aeruginosa virulence drug development by targeting the AQ receptor PqsR

Unfulfilled farmer expectations: the case of the Insect Resistant Maize for Africa (IRMA) project in Kenya

<p>Abstract</p> <p>Background</p> <p>Maize is the most important staple food in Kenya; any reduction in production and yield therefore often becomes a national food security concern. To address the challenge posed by the maize stem borer, the Insect Resistant Maize for Africa (IRMA) agricultural biotechnology public-private partnership (PPP) project was launched in 1999. There were, however, pre-existing concerns regarding the use of genetic engineering in crop production and skepticism about private sector involvement. The purpose of this case study was to understand the role of trust in the IRMA partnership by identifying the challenges to, and practices for, building trust in the project.</p> <p>Methods</p> <p>Data were collected by conducting face-to-face, semi-structured interviews; reviewing publicly available project documents; and direct observations. The data were analyzed to generate recurring and emergent themes on how trust is understood and built among the partners in the IRMA project and between the project and the community.</p> <p>Results</p> <p>Clear and continued communication with stakeholders is of paramount importance to building trust, especially regarding competition among partners about project management positions; a lack of clarity on ownership of intellectual property rights (IPRs); and the influence of anti-genetic modification (GM) organizations. Awareness creation about IRMA’s anticipated products raised the end users’ expectations, which were unfulfilled due to failure to deliver <it>Bacillus thuringiensis</it> (Bt)-based products, thereby leading to diminished trust between the project and the community.</p> <p>Conclusions</p> <p>Four key issues have been identified from the results of the study. First, the inability to deliver the intended products to the end user diminished stakeholders’ trust and interest in the project. Second, full and honest disclosure of information by partners when entering into project agreements is crucial to ensuring progress in a project. Third, engaging stakeholders and creating awareness immediately at the project’s inception contributes to trust building. Fourth, public sector goodwill combined with private sector technology and skills are necessary for a successful partnership. These findings may serve as a useful guide for building and fostering trust among partners in other agbiotech PPPs in sub-Saharan Africa.</p

Harmonized biosafety regulations are key to trust building in regional agbiotech partnerships: the case of the Bt cotton project in East Africa

<p>Abstract</p> <p>Background</p> <p>The <it>Bacillus thuringiensis</it> (Bt) cotton public-private partnership (PPP) project in East Africa was designed to gather baseline data on the effect of Bt cotton on biodiversity and the possibility of gene flow to wild cotton varieties. The results of the project are intended to be useful for Kenya, Uganda, and Tanzania when applying for biosafety approvals. Using the backdrop of the different biosafety regulations in the three countries, we investigate the role of trust in the Bt cotton partnership in East Africa.</p> <p>Methods</p> <p>Data were collected by reviewing relevant project documents and peer-reviewed articles on Bt cotton in Tanzania, Kenya and Uganda; conducting face-to-face interviews with key informants of the project; and conducting direct observations of the project. Data were analyzed based on recurring and emergent themes to create a comprehensive narrative on how trust is understood and built among the partners and with the community.</p> <p>Results</p> <p>We identified three factors that posed challenges to building trust in the Bt cotton project in East Africa: different regulatory regimes among the three countries; structural and management differences among the three partner institutions; and poor public awareness of GM crops and negative perceptions of the private sector. The structural and management differences were said to be addressed through joint planning, harmonization of research protocols, and management practices, while poor public awareness of GM crops and negative perceptions of the private sector were said to be addressed through open communication, sharing of resources, direct stakeholder engagement and awareness creation. The regulatory differences remained outside the scope of the project.</p> <p>Conclusions</p> <p>To improve the effectiveness of agbiotech PPPs, there is first a need for a regulatory regime that is acceptable to both the public and private sector partners. Second, early and continuous joint planning; sharing of information; and transparency encourages accountability and fosters trust building. Third, direct stakeholder engagement and awareness creation builds trust between project partners and the community. A concern raised by the interviewees was the absence of a regulatory framework in Tanzania, which deterred active private sector participation in the project.</p