Novel Viability Loss Process Induced by Electric Fields is Observed in the Extremophilic Deinococcus Radiodurans Exposed to Gamma Radiation

Abstract

Previously, we proposed a novel mechanism of antibacterial translocation through the general diffusion porin OmpC in Gram-negative E. coli (Abstract 12-A-3857-BPS, 2668-Pos). Our proposed mechanism explains the observed rapid translocation of substrates capable of replacing the H-bonds of solvating water in the channel constriction zone upon evacuation of this water to bulk solvent. This hypothesis is based on previous work linking protein-ligand association and dissociation barriers in general to the costs of water transfer to/from binding sites. In the current study, we investigated a set of 8 carbapenems (including known drugs and antibacterials in development) for their structure-translocation kinetic relationships with OmpC using electrophysiology and microbiology methods. Taking into account the supporting electrophysiology and microbiology data, we demonstrated that these molecules likewise translocate through OmpC porin at rates that depend on their polar composition and H-bond replacement ability. The additional acidic group of ertapenem compared to other analogs promoted the highest entry rate into OmpC (kon~2x104 M-1s-1). Zwitterionic compounds with highly polar groups attached to the penem-2 ring (i.e. panipenem, imipenem and doripenem) exhibited faster kon (>104 M-1s-1), while those zwitterionic analogs with fewer exposed polar groups (i.e. meropenem and biapenem) exhibited slower kon (~5x103 M-1s-1). Tebipenem pivoxil, a pro-dug developed for oral absorption, and razupenem with a change to thiazol-2-yl-thio moiety exhibited the slowest kon rates in to OmpC (~1.5x103 M-1s-1) and also showed interaction with the phospholipid membrane. Our findings may help better understand the molecular mechanisms underlying antibiotic uptake through the outer membrane of Gram-negative bacteria, which is a key step in achieving antibiotic exposure and efficacy at intracellular targets.

Sensing Of DNA Damage, Instantly Activation of Repairing Proteins and Radio Sensitizers-A Biophysical Model

“…A series of experiments was carried out at this Laboratory with prokaryotes and eukaryotes exposed to ionizing radiation and exogenous static electric fields. [22][23][24][25] All results show that when cells exposed to radiation are immediately submitted to static electric fields, cell death increases enormously compared to the effect of radiation alone. Since the sole application of an electric (E) field has no effect on control samples, it was thus demonstrated that the E-field acts on the DNA repair process.…”

“…The limits of this radio sensitization process induced by SEFs were tested by carrying out viability measurements with highly radioresistant cell lines, such as Deinococcus radiodurans, one of the fiercest radioresistant organisms, exhibiting a sophisticated repair mechanism responsible for its extremophilic character. 22,24 It should be noted that the present approach, using long-range electron transfer and conduction in DNA (Appendix 1) is high on the agenda of many biochemistry and biophysics research groups, as evidenced by a Special Issue section in PNAS-102-see, in particular. [30][31][32] The hypothesis here developed, by constituting a reasonable guideline for more elaborated signaling/transduction models, is also an original and effective contribution to the research field on cellular responses to genomic insult.…”

Desenvolvimento de um modelo biofísico para a descrição do processo de reparo de quebra-dupla em DNA

“…). Detalhes em (Arruda-Neto et al, 2014) Concluindo esta análise, os resultados evidenciam que a aplicação de campos elétricos estáticos (CEE) em células anteriormente expostas à radiação ionizante incrementa grandemente a morte celular. O efeito é similar ao observado quando uma radiação muito ionizante (LET grande) é usada (Han & Yu, 2009;Ball, Moore & Turner, 2008;Attar, Molaje & Khansari, 2007;Mozumder & Hatano, 2004;Ritter, Cleaver & Tobias, 1977).…”

“…não seria suficiente para polarizar a proteína. Contudo, resultados experimentais mostram que ocorreu polarização de proteínas de reparo (videArruda-Neto et al, 2014;Arruda- Neto et al, 2010 e Arruda-Neto et al, 2009. Portanto, algum ingrediente de natureza física, responsável pela produção de um E  bem maior, foi ignorado.…”

“…As proteínas de reparo seriam polarizadas na direção do CCE aplicado, via rotação dos momentos de dipolo daquelas pelo torque aplicado pelo campo elétrico exógeno. Essa polarização, por sua vez constitui uma interferência negativa nos mecanismos de reparo (detalhes emArruda-Neto et al, 2014;Arruda-Neto et al, 2010;Arruda-Neto et al, 2009).…”