Theoretical study of the hydroxylation of phenols mediated by an end-on bound superoxo–copper(II) complex

Peptidylglycine α-amidating monooxygenase and dopamine β-monooxygenase are copper-containing proteins which catalyze essential hydroxylation reactions in biological systems. There are several possible mechanisms for the reductive O₂-activation at their mononuclear copper active site. Recently, Karlin and coworkers reported on the reactivity of a copper(II)–superoxo complex which is capable of inducing the hydroxylation of phenols with incorporated oxygen atoms derived from the Cu(II)-O₂ ^·− moiety. In the present work the reaction mechanism for the abovementioned superoxo complex with phenols is studied. The pathways found are analyzed with the aim of providing some insight into the nature of the chemical and biological copper-promoted oxidative processes with 1:1 Cu(I)/O₂-derived species. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Theoretical study of cyclohexane hydroxylation by three possible isomers of [FeIV(O)(R-TPEN)]2+: does the pentadentate ligand wrapping around the metal center differently lead to the different stability and reactivity?

Density functional theory calculations have been carried out to elucidate the mechanism of cyclohexane hydroxylation by three possible isomers of [Fe^IV(O)(N-R-N,N′,N′-tris(2-pyridylmethyl)ethane-1,2-diamine)]²⁺ (R is methyl or benzyl) (Klinker et al. in Angew Chem Int Ed 44:3690–3694, 2005). The calculations offer a mechanistic view and reveal the following features: (a) all the three isomers possess triplet ground states and low-lying quintet excited states, (b) the relative stability follows the order isomer A > isomer B > isomer C, in agreement with the conclusions of Klinker et al., (c) the theoretical investigations provide a rationale to explain the interconversion of the three isomers, (d) the reaction pathways of the C–H hydroxylation are initiated by a hydrogen-abstraction step, and (e) the three isomers react with cyclohexane via two-state-reactivity patterns on competing triplet and quintet spin-state surfaces. As such, in the gas phase, the relative reactivity exhibits the trend isomer B > isomer A, while at the highest level, B2//B1 with zero point energy and solvation corrections, the relative reactivity follows the order isomer B > isomer A > isomer C. Thus, the calculated reaction pathway shows that pyridine rings perpendicular to the Fe–O axis result in more reactive species, and a pyridine ring coordinated trans to the oxygen atom leads to the least reactive isomer. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.

Theoretical study of the hydroxylation of phenolates by the Cu2O2(N,N′-dimethylethylenediamine)2 2+ complex

Tyrosinase catalyzes the ortho hydroxylation of monophenols and the subsequent oxidation of the diphenolic products to the resulting quinones. In efforts to create biomimetic copper complexes that can oxidize C–H bonds, Stack and coworkers recently reported a synthetic μ-η²:η²-peroxodicopper(II)(DBED)₂ complex (DBED is N,N′-di-tert-butylethylenediamine), which rapidly hydroxylates phenolates. A reactive intermediate consistent with a bis-μ-oxo-dicopper(III)-phenolate complex, with the O–O bond fully cleaved, is observed experimentally. Overall, the evidence for sequential O–O bond cleavage and C–O bond formation in this synthetic complex suggests an alternative mechanism to the concerted or late-stage O–O bond scission generally accepted for the phenol hydroxylation reaction performed by tyrosinase. In this work, the reaction mechanism of this peroxodicopper(II) complex was studied with hybrid density functional methods by replacing DBED in the μ-η²:η²-peroxodicopper(II)(DBED)₂ complex by N,N′-dimethylethylenediamine ligands to reduce the computational costs. The reaction mechanism obtained is compared with the existing proposals for the catalytic ortho hydroxylation of monophenol and the subsequent oxidation of the diphenolic product to the resulting quinone with the aim of gaining some understanding about the copper-promoted oxidation processes mediated by 2:1 Cu(I)O₂-derived species. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Microbial community of acetate utilizing denitrifiers in aerobic granules

Nitrite accumulates during biological denitrification processes when carbon sources are insufficient. Acetate, methanol, and ethanol were investigated as supplementary carbon sources in the nitrite denitrification process using biogranules. Without supplementary external electron donors (control), the biogranules degraded 200 mg l⁻¹ nitrite at a rate of 0.27 mg NO₂–N g⁻¹ VSS h⁻¹. Notably, 1,500 mg l⁻¹ acetate and 700 mg l⁻¹ methanol or ethanol enhanced denitrification rates for 200 mg l⁻¹ nitrite at 2.07, 1.20, and 1.60 mg NO₂–N g⁻¹ VSS h⁻¹, respectively; these rates were significantly higher than that of the control. The sodium dodecyl sulfate polyacrylamide gel electrophoresis of the nitrite reductase (NiR) enzyme identified three prominent bands with molecular weights of 37–41 kDa. A linear correlation existed between incremental denitrification rates and incremental activity of the NiR enzyme. The NiR enzyme activity was enhanced by the supplementary carbon sources, thereby increasing the nitrite denitrification rate. The capacity of supplementary carbon source on enhancing NiR enzyme activity follows: methanol > acetate > ethanol on molar basis or acetate > ethanol > methanol on an added weight basis.     

Macrobenthic fauna of the Franz Josef Land archipelago

Benthic fauna of the deeper waters (50–320 m) of the Franz Josef Land archipelago sampled in August 1992 using standard western techniques is described and compared with earlier Russian studies of the archipelago. Three faunal assemblages are recognised at depth ranges between 50–150, 150–300, and >300 m depth. Their distributions are related to depth and to different water masses derived from a mixture of Barents Sea water entering from the south at intermediate depths (20–150 m), and Atlantic water from the north below 300 m. The recorded faunal diversity and biomass is as high as or higher than that from comparable Arctic and northern boreal areas. Comparisons are also made with a complimentary Russian survey in the same area using different techniques. The Russian work collected larger organisms most effectively, whereas the present work recorded the smaller taxa more efficiently. Russian surveys provide complementary data to western studies, but care must be taken when comparing studies as they reflect biases in sampling gear and scientific emphasis. These data intercalibrations are important as Russian archives contain a wealth of information that, if used correctly in association with western data sources, will be useful in environmental monitoring studies as human activities increase in the Eurasian Arctic. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Biotransformation of menthol and geraniol by hairy root cultures of <Emphasis Type="Italic">Anethum graveolens</Emphasis>: effect on growth and volatile components

Two oxygen-containing monoterpene substrates, menthol or geraniol (25 mg l⁻¹), were added to Anethum graveolens hairy root cultures to evaluate the influence of the biotransformation capacity on growth and production of volatile compounds. Growth was assessed by the dissimilation method and by fresh and dry weight measurement. The volatiles were analyzed by GC and GC–MS. The total constitutive volatile component was composed, in more than 50%, by falcarinol (17–52%), apiole (11–24%), palmitic acid (7–16%), linoleic acid (4–9%), myristicin (4-8%) and n-octanal (2-5%). Substrate addition had no negative influence on growth. The relative amount of menthol quickly decreased 48 h after addition, and the biotransformation product menthyl acetate was concomitantly formed. Likewise, the added geraniol quickly decreased over 48 h alongside with the production of the biotransformation products. The added geraniol was biotransformed in 10 new products, the alcohols linalool, α-terpineol and citronellol, the aldehydes neral and geranial, the esters citronellyl, neryl and geranyl acetates and linalool and nerol oxides. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Effect of ultrafine-grained titanium surfaces on adhesion of bacteria

The influence of the ultrafine crystallinity of commercial purity grade 2 (as-received) titanium and titanium modified by equal channel angular pressing (modified titanium) on bacterial attachment was studied. A topographic profile analysis of the surface of the modified titanium revealed a complex morphology of the surface. Its prominent micro- and nano-scale features were 100–200-nm-scale undulations with 10–15 μm spacing. The undulating surfaces were nano-smooth, with height variations not exceeding 5–10 nm. These surface topography characteristics were distinctly different from those of the as-received samples, where broad valleys (up to 40–60 μm) were detected, whose inner surfaces exhibited asperities approximately 100 nm in height spaced at 1–2 μm. It was found that each of the three bacteria strains used in this study as adsorbates, viz. Staphylococcus aureus CIP 68.5, Pseudomonas aeruginosa ATCC 9025 and Escherichia coli K12, responded differently to the two types of titanium surfaces. Extreme grain refinement by ECAP resulted in substantially increased numbers of cells attached to the surface compared to as-received titanium. This enhanced degree of attachment was accompanied with an increased level of extracellular polymeric substances (EPS) production by the bacteria. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Impacts of deep open drains on water quality and biodiversity of receiving waterways in the Wheatbelt of Western Australia

Extensive networks of deep drains are being built in Western Australia to reduce the effects of dryland salinity on agricultural lands. Most of these drains discharge into natural river and wetland systems, with little consideration given to the environmental impacts. This study examined the downstream ecological impacts of one of the oldest deep drain networks in Western Australia, located in the Wakeman subcatchment near Narembeen. Twelve sites were sampled bi-monthly from October 2004 to September 2006. On each occasion, water quality parameters were measured and the macro-invertebrate fauna was sampled. Significant differences in water quality and macro-invertebrates were observed between the untreated sites and those affected by the drain discharge. Surface water at untreated sites was always fresh (<3 ppt), alkaline (pH 7.6–8.9) and turbid (49–600 NTU), whereas treatment sites were always saline (28–147 ppt), acidic (pH 1.9–3.8) and mostly clear (0–100 NTU). No recovery of water quality was observed with distance from discharge point (20 km). Invertebrates reflected differences in water quality, with drain discharge resulting in a sharp decline in species richness, and significant changes in macro-invertebrate community composition. Sites affected by drain discharge were dominated by fly larvae such as Orthocladiinae and Ceratopogonidae. Microcrustaceans were far more abundant at sites unaffected by drainage. The ecological values of Wheatbelt streams are likely to be further compromised by discharge of poor water quality from deep drainage. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Handling editor: K. Martens     

Deuterium isotope effects on <Superscript>15</Superscript>N backbone chemical shifts in proteins

Quantum mechanical calculations are presented that predict that one-bond deuterium isotope effects on the ¹⁵N chemical shift of backbone amides of proteins, ¹Δ¹⁵N(D), are sensitive to backbone conformation and hydrogen bonding. A quantitative empirical model for ¹Δ¹⁵N(D) including the backbone dihedral angles, Φ and Ψ, and the hydrogen bonding geometry is presented for glycine and amino acid residues with aliphatic side chains. The effect of hydrogen bonding is rationalized in part as an electric-field effect on the first derivative of the nuclear shielding with respect to N–H bond length. Another contributing factor is the effect of increased anharmonicity of the N–H stretching vibrational state upon hydrogen bonding, which results in an altered N–H/N–D equilibrium bond length ratio. The N–H stretching anharmonicity contribution falls off with the cosine of the N–H···O bond angle. For residues with uncharged side chains a very good prediction of isotope effects can be made. Thus, for proteins with known secondary structures, ¹Δ¹⁵N(D) can provide insights into hydrogen bonding geometries. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Cloning and expression of three peptide-linked β2-microglobulin molecules in <Emphasis Type="Italic">Escherichia coli</Emphasis> with an isocaudamer technique

The application of the peptide-linked β2-microglobulin (β2m) strategy is limited in some cases due to the incompatibility between the sequences of the peptides and the restriction sites of the plasmid vectors. An isocaudamer technique was adapted to overcome this restriction. Three peptide-linked β2m genes, HBc_18–27-hβ2m gene, OVA_257–264-mβ2m gene and HER2/neu_369–377-mβ2m gene, were inserted into the pET28a vectors with this technique. The corresponding proteins were expressed in Escherichia coli with yields of over 50 mg/l culture and purities of over 80%. This strategy facilitates the construction of peptide-linked β2m molecules and will simplify the preparation of major histocompatibility complex-peptide complexes. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Targeted Herceptin–dextran iron oxide nanoparticles for noninvasive imaging of HER2/neu receptors using MRI

A novel magnetic resonance imaging (MRI) contrast agent containing Herceptin is reported. The surfaces of superparamagnetic iron oxide nanoparticles were modified with dextran and conjugated with Herceptin (Herceptin–nanoparticles) to improve their dispersion, magnetization, and targeting of the specific receptors on cells. From analytical results, we found that Herceptin–nanoparticles were well dispersed in solutions of various pH range, and had no hysteresis, high saturation magnetization (80 emu/g), and low cytotoxicity to a variety of cells. Notably, the magnetic resonance enhancements for the different breast cancer cell lines (BT-474, SKBR-3, MDA-MB-231, and MCF-7) are proportional to the HER2/neu expression level in vitro. When Herceptin–nanoparticles were administered to mice bearing breast tumor allograft by intravenous injection, the tumor site was detected in T ₂-weighted magnetic resonance images as a 45% enhancement drop, indicating a high level of accumulation of the contrast agent within the tumor sites. Therefore, targeting of cancer cells was observed by in vitro and in vivo MRI studies using Herceptin–nanoparticles contrast agent. In addition, Herceptin–nanoparticles enhancing the magnetic resonance signal intensity were sufficient to detect the cell lines with a low level of HER2/neu expression. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Spatial behaviour of little owls (Athene noctua) in a declining low-density population in Denmark

Knowledge of a species’ spatial behaviour is essential for understanding its behavioural ecology, as well as a prerequisite to planning of conservation strategies. The Little Owl (Athene noctua) has shown a substantial decline in north-western Europe and is on the road of extinction in Denmark. To quantify relevant aspects of spatial behaviour in the last remaining Danish population, we followed 27 radio-tagged owls representing 14 territories during a period of 2 years. Mated owls were resident at nesting sites year-round with half of all nocturnal locations found within 125 m. Mean nightly distance from roosts peaked in January at 249 m (95% CI = 195–319) and dipped in May to 89 m (66–121). Distance from roosts varied non-linearly with temperature with the longest distances found at 6°C. Home range size varied with more than a tenfold difference between pairs, with pairs with neighbours maintaining 2–3 times larger ranges than isolated pairs independent of habitat composition. Mean home range size of 14 pairs was 41 (95% CI 27–64) and 2.6 ha (1.5–4.5) for 90 and 50% minimum convex polygons, respectively. Males and females behaved similarly, except in the breeding season where females foraged closer to the nest. Mates overlapped completely in activity distributions and were located closer to each other than expected by chance, suggesting a permanent pair-bond. Inter-mate distances were not shorter prior to egg laying compared to the rest of the year, indicating little male investment in mate guarding. No mates left their partner, but widowed birds left their territory within 6–12 months if unable to attract a new mate. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Ecological relationships between phytoplankton communities and different spatial scales in European reservoirs: implications at catchment level monitoring programmes

Phytoplankton communities are structured by factors acting over temporal and spatial scales. Identifying which factors are driving spatial patterns in aquatic communities is the central aim of ecology. In this study, data sets of phytoplankton communities and environmental data of two Portuguese reservoirs types (lowland “riverine reservoirs” and higher altitude “artificial lake reservoirs”) were used to determine the importance of environmental variables at different spatial (geographical, regional and local) and time scales (seasons, years) on the community structure. In all the data sets, the multivariate ordination technique Canonical Correspondence Analysis (CCA) showed that regional and local scales explained the majority (9–18% and 13–19%, respectively) of the taxa variance. However, for “riverine reservoirs”, time variables were more important, explaining 27% of the variability in phytoplankton assemblages. Variance partitioning was used to assess the individual importance of the three spatial scales and time for the community structure of the two reservoir types. The majority of among-site variability (5.9–21.4%) was accounted for by time variables, with local, regional, and geographical scale variables accounting for 3.3–5.6%, 3.7–4.5% and 2.6–2.9%, respectively. The effects of different spatial scales on phytoplankton communities were clearly interrelated; thus, implying that phytoplankton assemblages are capable of detecting stress from catchment to site scales. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Handling editor: J. Padisak     

The molecular structure of a curl-shaped retinal isomer

Computational studies of retinal protonated Schiff base (PSB) isomers show that a twisted curl-shaped conformation of the retinyl chain is a new low-lying minimum on the ground-state potential energy surface. The curl-shaped isomer has a twisted structure in the vicinity of the C₁₁=C₁₂ double bond where the 11-cis retinal PSB isomerizes in the rhodopsin photoreaction. The twisted configuration is a trapped structure between the 11-cis and all-trans isomers. Rotation around the C₁₀–C₁₁ single bond towards the 11-cis structure is prevented by steric interactions of the two methyl groups on the retinyl chain and by the torsion barrier of the C₁₀–C₁₁ bond in the other direction. Calculations of spectroscopic properties of the 11-cis, all-trans, and curl-shaped isomers provide useful data for future identification of the new retinal PSB isomer. Circular dichroism (CD) spectroscopy might be used to distinguish between the retinal PSB isomers. The potential energy surface for the orientation of the β-ionone ring of the 11-cis retinal PSB reveals three minima depending on the torsion angle of the β-ionone ring. Two of the minima correspond to 6-s-cis configurations and one has the β-ionone ring in 6-s-trans position. The calculated CD spectra for the two 6-s-cis configurations differ significantly indicating that the sign of the β-ionone ring torsion angle could be determined using CD spectroscopy. Calculations of the CD spectra suggest that a flip of the β-ionone ring might occur during the first 1 ps of the photoreaction. Rhodopsin has a negative torsion angle for the β-ionone ring, whereas the change in the sign of the first peak in the experimental CD spectrum for bathorhodopsin could suggest that it has a positive torsion angle for the β-ionone ring. Calculated nuclear magnetic resonance (NMR) shielding constants and infrared (IR) spectra are also reported for the retinal PSB isomers. Figure The figure shows the optimized molecular structure of the curl-shaped retinal isomer. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Phylogenetic position of the monotypic Des Murs’ Wiretail ( Sylviorthorhynchus desmursii , Aves: Furnariidae) based on mitochondrial and nuclear DNA

Parallelisms and paraphyletic assemblages are common among ovenbirds. Molecular markers are therefore the best approach when studying the evolutionary relationships among the members of this unparalleled diversified family. We obtained nucleotide sequence data from mitochondrial (cytochrome b) and nuclear genes (myoglobin and glyceraldehyde-3-phosphodehydrogenase) and used these to deduce the phylogenetic position of a monotypic genus endemic to the austral temperate rainforests of southern South America, the Des Murs’ Wiretail (Sylviorthorhynchus desmursii Des Murs, 1847, Aves: Furnariidae). Phylogenetic analyses based on maximum parsimony, maximum likelihood and Bayesian inference all converged into a congruent topology, with a basal position of Des Murs’ Wiretail within Synallaxinae together with Tit–Spinetails (Leptasthenura). Our data reject the hypothesis of a phylogenetic relationship between Des Murs’ Wiretail and thistletails (Schizoeaca) which exhibit parallelisms in morphology, tail structure and nest architecture. Using a molecular clock based on the myoglobin intron 2 gene, we estimated a divergence time of Des Murs’ Wiretail from Tit-Spinetails of 14–15 Myr, which is associated with the appearance of sclerophyllous forest elements in Chile at the Middle–Upper Miocene. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Arecoline and the 30–100 kDa fraction of areca nut extract differentially regulate mTOR and respectively induce apoptosis and autophagy: a pilot study

Areca nut (AN) is recognized as a human carcinogen; however, few studies of the cytotoxic effects of AN ingredients on cells have been reported. In Taiwan, AN, lime and inflorescence of Piper betle are the common components of betel quid (BQ). We recently noticed that extract of AN (ANE), but not those of lime and inflorescence of Piper betle, induces rounding cell morphology and nuclear shrinkage in different types of carcinoma cells. In this study, the rounding cell activity was first traced to the partially purified ≥10 kDa fraction (ANE ≥ 10 K) and subsequently to the 30–100 kDa fraction (ANE 30–100 K). ANE and ANE ≥10 K stimulated nuclear shrinkage (P < 0.001 in both cases) and the clearance of the cytoplasm. ANE, ANE ≥ 10 K, and ANE 30–100 K induced the cleavage of LC3-I (P < 0.05, 0.01, and 0.05, respectively) and the emergence of autophagic vacuoles (AVs) and acidic vesicles. On the other hand, arecoline (Are, the major alkaloid of AN) triggered caspase-3 activation, peri-nuclear chromatin condensation, and micronucleation. Meanwhile, ANE 30–100 K, but not Are, inhibited the phosphorylation of the mammalian target of rapamycin (mTOR)-Ser²⁴⁴⁸. In conclusion, this study demonstrates that different AN ingredients exerting differential impact on mTOR-Ser²⁴⁴⁸ phosphorylation are capable of triggering apoptosis and autophagy. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Shyun-Yeu Liu, Mei-Huei Lin, Yu-Rung Hsu and Ya-Yun Shih contributed equally to this work.     

An interrupted beta-propeller and protein disorder: structural bioinformatics insights into the N-terminus of alsin

Defects in the human ALS2 gene, which encodes the 1,657-amino-acid residue protein alsin, are linked to several related motor neuron diseases. We created a structural model for the N-terminal 690-residue region of alsin through comparative modelling based on regulator of chromosome condensation 1 (RCC1). We propose that this alsin region contains seven RCC1-like repeats in a seven-bladed beta-propeller structure. The propeller is formed by a double clasp arrangement containing two segments (residues 1–218 and residues 525–690). The 306-residue insert region, predicted to lie within blade 5 and to be largely disordered, is poorly conserved across species. Surface patches of evolutionary conservation probably indicate locations of binding sites. Both disease-causing missense mutations—Cys157Tyr and Gly540Glu—are buried in the propeller and likely to be structurally disruptive. This study aids design of experimental studies by highlighting the importance of construct length, will enhance interpretation of protein–protein interactions, and enable rational site-directed mutagenesis. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Litre-scale microbial fuel cells operated in a complete loop

Using the anode effluent to compensate the alkalinization in a bio-cathode has recently been proposed as a way to operate a microbial fuel cell (MFC) in a continuous and pH neutral way. In this research, we successfully demonstrated that the operation of a MFC without any pH adjustments is possible by completing the liquid loop over cathode and anode. During the complete loop operation, a stable current production of 23.2 ± 2.5 A m⁻³ MFC was obtained, even in the presence of 3.2–5.2 mg O₂ L⁻¹ in the anode. The use of current collectors and subdivided electrical circuitries for relative large 2.5-L-scale MFCs resulted in ohmic cell resistances in the order of 1.4–1.7 mΩ m³ MFC, which were comparable to values of ten times smaller MFCs. Nevertheless, the bio-cathode activity still needs to be improved significantly with a factor 10–50 in order achieve desirable current densities of 1,000 A m⁻³ MFC. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Regulation of the Kv2.1 Potassium Channel by MinK and MiRP1

Kv2.1 is a voltage-gated potassium (Kv) channel α-subunit expressed in mammalian heart and brain. MinK-related peptides (MiRPs), encoded by KCNE genes, are single–transmembrane domain ancillary subunits that form complexes with Kv channel α-subunits to modify their function. Mutations in human MinK (KCNE1) and MiRP1 (KCNE2) are associated with inherited and acquired forms of long QT syndrome (LQTS). Here, coimmunoprecipitations from rat heart tissue suggested that both MinK and MiRP1 form native cardiac complexes with Kv2.1. In whole-cell voltage-clamp studies of subunits expressed in CHO cells, rat MinK and MiRP1 reduced Kv2.1 current density three- and twofold, respectively; slowed Kv2.1 activation (at +60 mV) two- and threefold, respectively; and slowed Kv2.1 deactivation less than twofold. Human MinK slowed Kv2.1 activation 25%, while human MiRP1 slowed Kv2.1 activation and deactivation twofold. Inherited mutations in human MinK and MiRP1, previously associated with LQTS, were also evaluated. D76N–MinK and S74L–MinK reduced Kv2.1 current density (threefold and 40%, respectively) and slowed deactivation (60% and 80%, respectively). Compared to wild-type human MiRP1–Kv2.1 complexes, channels formed with M54T– or I57T–MiRP1 showed greatly slowed activation (tenfold and fivefold, respectively). The data broaden the potential roles of MinK and MiRP1 in cardiac physiology and support the possibility that inherited mutations in either subunit could contribute to cardiac arrhythmia by multiple mechanisms. Electronic supplementary material  The online version of this article (doi: ) contains supplementary material, which is available to authorized users. Z. A. McCrossan and T. K. Roepke have contributed equally to this work.