A thermostable quorum-quenching lactonase from Geobacillus kaustophilus HTA426 (GI: 56420041) was used as an initial template for in vitro directed evolution experiments. This enzyme belongs to the phosphotriesterase-like lactonase (PLL) group of enzymes within the amidohydrolase superfamily that hydrolyze N-acylhomoserine lactones (AHLs) that are involved in virulence pathways of quorum-sensing pathogenic bacteria. Here we have determined the N-butyryl-L-homoserine lactone-liganded structure of the catalytically inactive D266N mutant of this enzyme to a resolution of 1.6 Å . Using a tunable, bioluminescence-based quorum-quenching molecular circuit, the catalytic efficiency was enhanced, and the AHL substrate range increased through two point mutations on the loops at the C-terminal ends of the third and seventh -strands. This E101N/R230I mutant had an increased value of k cat /K m of 72-fold toward 3-oxo-N-dodecanoyl-L-homoserine lactone. The evolved mutant also exhibited lactonase activity toward N-butyryl-L-homoserine lactone, an AHL that was previously not hydrolyzed by the wild-type enzyme. Both the purified wild-type and mutant enzymes contain a mixture of zinc and iron and are colored purple and brown, respectively, at high concentrations. The origin of this coloration is suggested to be because of a charge transfer complex involving the -cation and Tyr-99 within the enzyme active site. Modulation of the charge transfer complex alters the lactonase activity of the mutant enzymes and is reflected in enzyme coloration changes. We attribute the observed enhancement in catalytic reactivity of the evolved enzyme to favorable modulations of the active site architecture toward productive geometries required for chemical catalysis.The amidohydrolase superfamily of enzymes comprises members that catalyze hydrolytic reactions on a broad range of substrates bearing ester or amide functional groups with carbon or phosphorus centers (1). These reactions are mediated by a conserved mononuclear or binuclear center within a (/␣) 8 -barrel structural scaffold (2) and are initiated by the activation of a water molecule for nucleophilic attack on an activated scissile bond of the substrate for subsequent hydrolysis. The amidohydrolase superfamily was first described by Holm and Sander in 1997 (3) and has since expanded to cover more than 30 reactions involving a diverse range of substrates (2, 4), including quorum-sensing N-acylhomoserine lactones (AHLs) 4 (5). We recently reported the directed evolution of a member of the amidohydrolase superfamily that hydrolyzes AHLs (6); this quorum-quenching enzyme is part of a group of divergently related enzymes within the superfamily, the phosphotriesterase-like lactonases (PLLs), which hydrolyze quorumsensing AHLs. Quorum-sensing is an integral part of microbial interaction and is responsible for virulence or pathogenicity of disease-causing bacteria (7). Modulation and perturbation of this quorum-sensing pathway has been demonstrated in principle to be an effective "anti-microb...
In XXZ chains with large enough interactions, spin transport can be significantly suppressed when the bias of the dissipative driving becomes large enough. This phenomenon of negative differential conductance is caused by the formation of two oppositely polarized ferromagnetic domains at the edges of the chain. Here, we show that this many-body effect, combined with a non-uniform magnetic field, can allow for a high degree of control of the spin current. In particular, by studying all of the possible shapes of local magnetic fields potentials, we find that a configuration in which the magnetic field points up for half of the chain and down for the other half, can result in giant spin-current rectification, for example, up to 108 for a system with only 8 spins. Our results show clear indications that the rectification can increase with the system size.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.