In nature as in biotechnology, light-oxygen-voltage photoreceptors perceive blue light to elicit spatiotemporally defined cellular responses. Photon absorption drives thioadduct formation between a conserved cysteine and the flavin chromophore. An equally conserved, proximal glutamine processes the resultant flavin protonation into downstream hydrogen-bond rearrangements. Here, we report that this glutamine, long deemed essential, is generally dispensable. In its absence, several light-oxygen-voltage receptors invariably retained productive, if often attenuated, signaling responses. Structures of a light-oxygen-voltage paradigm at around 1 Å resolution revealed highly similar light-induced conformational changes, irrespective of whether the glutamine is present. Naturally occurring, glutamine-deficient light-oxygen-voltage receptors likely serve as bona fide photoreceptors, as we showcase for a diguanylate cyclase. We propose that without the glutamine, water molecules transiently approach the chromophore and thus propagate flavin protonation downstream. Signaling without glutamine appears intrinsic to light-oxygen-voltage receptors, which pertains to biotechnological applications and suggests evolutionary descendance from redox-active flavoproteins.

The enzyme glucose-6-phosphate dehydrogenase (G6PDH) catalyzes the first step of the oxidative branch of the pentose phosphate pathway, which provides cells with NADPH, an essential cofactor for many biosynthetic pathways and antioxidizing enzymes. In Trypanosoma cruzi, the G6PDH has being pursued as a relevant target for the development of new drugs against Chagas disease. At present, the best characterized inhibitors of T. cruzi G6PDH are steroidal halogenated compounds derivatives from the mammalian hormone precursor dehydroepiandrosterone, which indeed are also good inhibitors of the human homologue enzyme. The lack of target selectivity might result in hemolytic side effects due to partial inhibition of human G6PDH in red blood cells. Moreover, the treatment of Chagas patients with steroidal drugs might also cause undesired androgenic side effects. Aiming to identify of new chemical classes of T. cruzi G6PDH inhibitors, we performed a target-based high-throughput screen campaign against a commercial library of diverse compounds. Novel TcG6PDH inhibitors were identified among thienopyrimidine and quinazolinone derivatives. Preliminary structure activity relationships for the identified hits are presented, including structural features that contribute for selectivity toward the parasite enzyme. Our results indicate that quinazolinones are promising hits that should be considered for further optimization.

Trypanosoma cruzi is the causative agent of Chagas disease. The lack of an efficient and safe treatment supports the research into novel metabolic targets, with the malic enzyme (ME) representing one such potential candidate. T. cruzi expresses a cytosolic (TcMEc) and a mitochondrial (TcMEm) ME isoform, with these activities functioning to generate NADPH, a key source of reducing equivalents that drives a range of anabolic and protective processes. To identify specific inhibitors that target TcMEs, two independent high-throughput screening strategies using a diversity library containing 30,000 compounds were employed. IC values of 262 molecules were determined for both TcMEs, as well as for three human ME isoforms, with the inhibitors clustered into six groups according to their chemical similarity. The most potent hits belonged to a sulfonamide group that specifically target TcMEc. Moreover, several selected inhibitors of both TcMEs showed a trypanocidal effect against the replicative forms of T. cruzi. The chemical diversity observed among those compounds that inhibit TcMEs activity emphasizes the druggability of these enzymes, with a sulfonamide-based subset of compounds readily able to block TcMEc function at a low nanomolar range.