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Inhibidores de la proteína de división celular bacteriana tsZ dirigidos al sitio de unión del nucleótido

AutorVergonos, A.
DirectorHuecas, Sonia
Palabras claveFtsZ
División celular bacteriana
Interacciones proteína-ligando
Ensamblaje de proteínas
Fecha de publicación2017
EditorCSIC - Centro de Investigaciones Biológicas (CIB)
Universidad Autónoma de Madrid
ResumenBinary fission of many prokaryotes as well as some eukaryotic organelles depends on the FtsZ protein, the most conserved bacterial cell division protein (Margolin, 2005). FtsZ shares structural homology with eukaryotic protein tubulin. FtsZ polymerizes at the future division site to form a ring-like structure, called the Z-ring that serves as a scaffold to recruit all other division proteins (Xiao and Goley, 2016). Numerous inhibitors of FtsZ have been described recently (Schaffner-Barbero et al., 2012), and for some of them the binding region in the protein still remains unclear. To date, only two small molecule binding sites in the protein have been identified: the binding site of PC190723 (derived from 3-methoxybenzamide), a validated FtsZ inhibitor that binds between the N- and Cterminal domains of FtsZ (Haydon et al., 2008); and the nucleotide (GTP) binding site at the top end of the FtsZ monomer, with the complete GTPase site formed by the Ndomain of one monomer and the C-domain of the next monomer in the protofilament (Oliva et al., 2004). Derivatives of GTP molecule with substitutions in carbon 8 have also been reported as potent inhibitors (KdMorphGTP = 0.7 ± 0.1 μM) that compete with GTP for its binding site in the protein (Lappchen et al., 2008; Marcelo et al., 2013). The natural compounds crysophaentin (Plaza et al., 2010) and hemicrysophaentin (Keffer et al., 2013) have also shown affinity for the GTP binding site in FtsZ. Recently, studies in our laboratory have demonstrated that small synthetic inhibitors with different chemical structure to GTP can compete with the natural ligand for its binding to FtsZ (KdUCM44 =0.7 ± 0.1 μM, MIC: 25 μM in B. subtilis, and 22 μM in S. aureus MRSA; KdUCM53 =0.8 ± 0.2 μM, MIC: 13 μM in B. subtilis, and 4.7 μM in S. aureus MRSA) (Ruiz-Avila et al., 2013). These compounds share a general structure of two gallate subunits bound to a central core of 1,3- naphtalene by ester groups. Based on this first approach, we have synthesized new compounds with a 3,5-biphenyl scaffold instead the naphthalene group as a central core; this modification improves the affinity (KdUCM81 = 0.5 ± 0.1 μM) and antibacterial activity (MIC UCM81: 5 μM in B. subtilis, and 7 μM in S. aureus MRSA) (Ruiz-Ávila et al., 2013). This thesis has also been focused on the identification and study of new small molecules coming from a GTP site-based virtual screening, employing commercial libraries in the ICM database (Totrov y Abagyan, 1997). We have found a compound (VS2.18) that binds to the nucleotide site of Bacillus subtilis FtsZ with an affinity of 104 ± 38 μM and modulates its polymerization. Based on this compound we conducted a ligand-based search that let us found new compounds (VS2.25, VS2.33) with improved affinity for the GTP binding site (around 2.5 – 10 μM); that also inhibit B. subtilis cell division and growth. These compounds produced mislocalization of the FtsZ in the bacterial cells, and this effect was correlated with a phenotypic profile of filamented cells, which become unable to divide. They show MIC values in B. subtilis around 20 – 50 μM, and their antibacterial activity has been confirmed in S. aureus MRSA. These compounds are a starting point of a multidisciplinary program of biochemistry, chemical biology and computational analysis in order to provide a structure-activity relationship to understand the mechanisms of action of these inhibitors that could help to develop new antibacterial agents.
Descripción197 p.-48 fig.-13 tab.
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