AcrB is the major multidrug exporter in Escherichia coli. Although several substrate-entrances have been identified, specificity of these various transport paths remains unclear. Here we present evidence for a substrate channel (channel 3) from central cavity trimer, which connected directly to deep pocket without first passing switch-loop and proximal . Planar aromatic cations, such as ethidium, prefer 3 channels 1 2. The efflux through increases by targeted mutations not competition with...

Abstract RamR is a transcriptional repressor of the gene-encoding RamA protein, which controls expression multidrug efflux system genes acrAB-tolC . an important multidrug-resistance factor, however, its structure and identity molecules to it responds have been unknown. Here, we report crystal in complex with multiple drugs, including berberine, violet, dequalinium, ethidium bromide rhodamine 6G. All compounds are found interact Phe155 RamR, each compound surrounded by different amino acid...

A negative phototransduction feedback in rods and cones is critical for the timely termination of their light responses extending function to a wide range intensities. The calcium mechanisms that modulate have been studied extensively. However, corresponding modulation enable terminate rapidly adapt bright light, properties our daytime vision, are still not understood. In cones, guanylyl cyclase potentially key step modulation. activity modulated by calcium-binding activating proteins (GCAP1...

Signal transduction in rod cells begins with photon absorption by rhodopsin and leads to the generation of an electrical response. The response profile is determined molecular properties phototransduction components. To examine how correlate rod-response profile, we have generated a knock-in mouse replaced its E122Q mutant, which exhibits different from those wild-type (WT) rhodopsin. Knock-in rods exhibited photosensitivity about 70% WT. Correspondingly, their single-photon had amplitude...

Rod and cone photoreceptor cells that are responsible for scotopic photopic vision, respectively, exhibit photoresponses different from each other contain similar phototransduction proteins with distinctive molecular properties. To investigate the contribution of properties visual pigments to responses cells, we have generated knock-in mice in which rod pigment (rhodopsin) was replaced mouse green-sensitive (mouse green). The green successfully transported outer segments, though expression...

Abstract During infection, Salmonella senses and responds to harsh environments within the host. Persistence in a bile-rich environment is important for infect small intestine or gallbladder multidrug efflux system AcrAB-TolC required bile resistance. The genes encoding this are mainly regulated by ramRA locus, which composed of divergently transcribed ramA ramR genes. acrAB tolC transcriptionally activated RamA, whose gene itself repressed RamR. RamR recognizes multiple drugs; however,...

ABSTRACT The AcrAB-TolC system in Escherichia coli is an intrinsic RND-type multidrug efflux transporter that functions as a tripartite complex of the inner membrane AcrB, outer channel TolC, and adaptor protein AcrA. Although crystal structures each component this have been elucidated, structure whole has not solved. available shown AcrB TolC function trimers, but number AcrA molecules now under debate. Disulfide chemical cross-linking experiments indicated stoichiometry AcrB-AcrA-TolC...

Significance Anurans are unique in possessing two types of rod photoreceptor cells, red and green rods. Red rods express rhodopsin, whereas blue-sensitive cone visual pigment. Rhodopsin exhibits a low rate thermal isomerization the retinal chromophore, which enables to detect photons with extremely high signal-to-noise for scotopic vision. Here, we show that anuran pigments acquired rhodopsin-like property through single amino acid mutation at position 47 evolutionary process from other...

Journal Article High-resolution Crystal Structure of β2-Microglobulin Formed at pH 7.0 Get access Kentaro Iwata, Iwata Institute for Protein Research, Osaka University, and CREST, Japan Science Technology Agency, Yamadaoka 3-2, Suita, 565-0871, Search other works by this author on: Oxford Academic PubMed Google Scholar Takanori Matsuura, Matsuura Kazumasa Sakurai, Sakurai Atsushi Nakagawa, Nakagawa Yuji Goto * *To whom correspondence should be addressed. Tel: +81-6-6879-8614, Fax:...

Cytochrome P450cam is a heme-containing enzyme which catalyzes hydroxylation of d-camphor. The heme bound in the pocket via noncovalent interactions, where two heme-propionate side chains interact with Arg, His, and/or Asp residues. To understand role heme-7-propionate chain, we prepared reconstituted an artificial one-legged has methyl group at position 7-propionate. Removal 7-propionate dramatically decreases d-camphor affinity by 3 orders magnitude relative to that wild-type enzyme, and...

Rod and cone photoreceptors use similar but distinct sets of phototransduction proteins to achieve different functional properties, suitable for their role as dim bright light receptors, respectively. For example, rod visual pigments couple variants the heterotrimeric G protein transducin. However, structural differences between transducin alpha subunits (Talpha) in determining rods cones is unknown. To address this question, we studied translocation signaling properties Talpha expressed...

Abstract Total synthesis of (−)‐enigmazole A, a marine macrolide natural product with cytotoxic activity, has been accomplished. The tetrahydropyran moiety was constructed by means domino olefin cross‐metathesis/intramolecular oxa‐Michael addition δ‐hydroxy olefin. After coupling advanced intermediates, the macrocycle formed through gold‐catalyzed rearrangement propargylic benzoate, followed ring‐closing metathesis resultant α,β‐unsaturated ketone.

Abstract RND-type multidrug efflux pumps have two voluminous multisite drug-binding pockets named the proximal and distal binding pocket. High- low-molecular-mass drugs bind to these pocket, respectively. Here, we report crystal structures of MexB Pseudomonas aeruginosa bound with high-molecular-mass compounds. Contrary expectations, lauryl maltose neopentyl glycol (LMNG, MW 1,005), which is a surfactant larger than pocket-binding drugs, was found pocket: one hydrophobic alkyl chains...

Calcium ions (Ca(2+)) modulate the phototransduction cascade of vertebrate cone photoreceptors to tune gain, inactivation, and light adaptation. In darkness, continuous current entering outer segment through cGMP-gated (CNG) channels is carried in part by Ca(2+), which then extruded back extracellular space. The mechanism Ca(2+) extrusion from mammalian cones not understood. dominant view has been that cone-specific isoform Na(+)/Ca(2+), K(+) exchanger, NCKX2, responsible for removing their...

Phosphodiesterase-6 (PDE6) is the key effector enzyme of vertebrate phototransduction pathway in rods and cones. Rod PDE6 catalytic core composed two distinct subunits, PDE6α PDE6β, whereas identical PDE6α′ subunits form cone core. It not known whether this difference subunit identity contributes to functional differences between To address question, we expressed photoreceptor cells retinal degeneration 10 (<i>rd10</i>) mouse that carries a mutation rod PDEβ subunit. We show adeno-associated...

The newt is an amazing four-limbed vertebrate that can regenerate various body parts including the retina. In this animal, when neural retina (NR) removed from eye by surgery (retinectomy), both NR and retinal pigment epithelium (RPE) eventually through process of reprogramming proliferation RPE cells. Thus far, we have pursued onset mechanism adult regeneration. study, using in vitro system, found mitogen-activated protein kinase (MAPK)/extracellular signal-regulated (ERK) (MEK)-ERK...

Abstract The newt, a group of urodele amphibians, has outstanding ability to repeatedly regenerate various body parts, even in the terrestrial life-stage. In this animal, when limb is amputated, cell mass named blastema appears on stump and eventually gives rise new functional limb. Erythrocytes (red blood cells) most non-mammalian vertebrates, including preserve their nucleus throughout life-span, although physiological roles such nucleated erythrocytes, other than oxygen delivery, are not...

Rpe65 −/− mice are unable to produce 11- cis -retinal, the chromophore of visual pigments. Consequently, pigment is present as apoprotein opsin with a minute level containing 9- -retinal chromophore. Notably, 10–20% fraction this mono-phosphorylated independently light conditions. To determine role rhodopsin kinase (GRK1) in phosphorylating and test whether eliminating phosphorylation would accelerate photoreceptor degeneration, we generated Grk1 mouse. The retinae had negligible...

Rod photoreceptors are exquisitely sensitive light detectors that function in dim light. The timely inactivation of their responses is critical for the ability rods to reliably detect and count photons. A key step rod transduction phosphorylation visual pigment, rhodopsin, catalyzed by G-protein-dependent receptor kinase 1 (GRK1). Absence GRK1 greatly prolongs photoreceptors' response enhances susceptibility degeneration. This study examined from mouse expressing various levels evaluate how...

Cytochrome P450cam (P450cam) binds a protoheme IX as prosthetic group via noncovalent interactions. Heme-6-propionate, one of the two heme-propionate side chains, forms hydrogen-bonding interactions with Arg112 and other hydrophilic amino acid residues. Here, we demonstrate structural functional roles 6-propionate chain in using reconstituted protein 6-depropionate-6-methylated (one-legged heme). The spectroscopic data enzymatic activities reveal that removal has no clear influence on enzyme...