Rapid Scan Mode Research Articles

Order-of-magnitude SNR improvement for high-field EPR spectrometers via 3D printed quasi-optical sample holders.

Here, we present a rapidly prototyped, cost-efficient, and 3D printed quasi-optical sample holder for improving the signal-to-noise ratio (SNR) in modern, resonator-free, and high-field electron paramagnetic resonance (HFEPR) spectrometers. Such spectrometers typically operate in induction mode: The detected EPR ("cross-polar") signal is polarized orthogonal to the incident ("co-polar") radiation. The sample holder makes use of an adjustable sample positioner that allows for optimizing the sample position to maximize the 240-gigahertz magnetic field B1 and a rooftop mirror that allows for small rotations of the microwave polarization to maximize the cross-polar signal and minimize the co-polar background. When optimally tuned, the sample holder was able to improve co-polar isolation by ≳20 decibels, which is proven beneficial for maximizing the SNR in rapid-scan, pulsed, and continuous-wave EPR experiments. In rapid-scan mode, the improved SNR enabled the recording of entire EPR spectra of a narrow-line radical in millisecond time scales, which, in turn, enabled real-time monitoring of a sample's evolving line shape.

Frequency chirped Fourier-Transform spectroscopy

Fast (sub-second) spectroscopy with high spectral resolution is of vital importance for revealing quantum chemistry kinetics of complex chemical and biological reactions. Fourier transform (FT) spectrometers can achieve high spectral resolution and operate at hundreds of ms time scales in rapid-scan mode. However, the linear translation of a scanning mirror imposes stringent time-resolution limitations to these systems, which makes simultaneous high spectral and temporal resolution very difficult. Here, we demonstrate an FT spectrometer whose operational principle is based on continuous rotational motion of the scanning mirror, effectively decoupling the spectral resolution from the temporal one. Furthermore, we show that such rotational FT spectrometer can perform Mid-IR dual-comb spectroscopy with a single comb source, since the Doppler-shifted version of the comb serves as the second comb. In our realization, we combine the advantages of dual-comb and FT spectroscopy using a single quantum cascade laser frequency comb emitting at 8.2 μm as a light source. Our technique does not require any diffractive or dispersive optical elements and hence preserve the Jacquinot’s-, Fellgett’s-, and Connes’-advantages of FT spectrometers. By integrating mulitple broadband sources, such system could pave the way for applications where high speed, large optical bandwidth, and high spectral resolution are desired.

Fabrication of bioactive poly(ether ether ketone) by laser melt infiltration of poly(ether ether ketone) inside the strontium apatite coatings

A versatile method to immobilize apatite nanoparticles onto PEEK substrate by using CO2 laser was demonstrated. Various strontium apatites were synthesized and dispersed in ethanol at a low concentration of citric acid. This dispersion of strontium apatites was coated on the surface of the PEEK substrate. The apatite-coated PEEK substrate was subsequently irradiated by CO2 laser with repeated multiple scanning mode. The surface temperature was monitored in real time and controlled at approximately the melting point of PEEK. The thermally fixed apatite coatings were unified with the PEEK substrate and unable to separate off. Dissolution of apatite layer in dilute acid solution revealed highly rugged surface morphologies of the PEEK substrate hidden inside the apatite coatings. From these observations we concluded that the heat-fixed apatite layer consisted of nanocomposite of apatite and PEEK. We propose a possible mechanism for the laser bonding processes of this apatite coating, that is, when the coating surface is irradiated by a CO2 laser with rapid periodic scanning mode, this irradiation initiates periodic temperature change with thermal expansion of the air on top of the apatite layer and may induce a photoacoustic effect on them, which may generate pressure waves to hold down the apatite matrix to the PEEK substrate. The heat on the apatite matrix also initiates melting of the adjacent PEEK surface and the melted PEEK polymer intrudes into the voids of the apatite matrix by capillary forces to produce nanocomposite of them. This surface-selective formation of PEEK/apatite nanocomposites on the PEEK substrate may be seen as an alternative method to the conventional surface coatings or various PEEK composite materials.The bioactivity of the apatite-coated PEEK substrate was affirmed through biomimetic deposition of calcium phosphate from simulated body fluid and dissolution behaviors of the apatite coatings in water was also evaluated.

On the geometric accuracy and stability of MSG SEVIRI images

Monitoring the geometric quality and temporal stability of Earth Observation satellite sensors is a major task of satellite vendors. As an owner and operator of widely used meteorological satellites, EUMETSAT (European Organisation for the Exploitation of Meteorological Satellites) carries out operational activities and scientific studies on the radiometric and geometric quality assessments of geostationary Meteosat satellites among others. As part of Level 1 Product Monitoring - Evolution Studies initiated by EUMETSAT, a new methodology has been developed and implemented as a prototype software, namely the Geometric Quality Assessment (GQA) Tool; which enhances image local texture, extracts prominent features and matches them via area-based least squares, and eliminates possible outliers using a set of rules. This article presents the results of algorithm validation activities carried out by using time series images of the Spinning Enhanced Visible and InfraRed Imager (SEVIRI) sensor aboard Meteosat Second Generation (MSG) satellites and MEdium Resolution Imaging Spectrometer (MERIS) global mosaics as reference to evaluate short- and long-term geometric temporal stability. Meteosat-11 SEVIRI time series data from a full-day and one year were employed for this purpose. In addition, the geometric quality of Meteosat-8 over Indian Ocean and Meteosat-10 in rapid scan mode was assessed for single acquisitions. The results show that the geometric accuracy of the evaluated datasets yield to the mean shifts up to one km in East-West (x) and North-South (y) directions, with larger errors in the y direction. The image inner geometric accuracy assessments revealed displacement errors with strip pattern in the three spectral bands (i.e. high-resolution visible, infra-red 10.8 and visible 0.8) evaluated within the framework.

Retrieval and Application of Rapid-Scan Atmospheric Motion Vectors Using an Infrared Channel of the INSAT-3DR Satellite

The advanced Indian meteorological satellite INSAT-3DR was configured in rapid-scan mode during the formation of tropical cyclone ‘Fani’ over the North Indian Ocean. This study is carried out to explore the possibility of further enhancement in atmospheric motion vectors (AMVs) to understand the extreme weather event using rapid-scan observations. A novel experiment has been performed to retrieve rapid-scan AMVs (RS-AMVs) with different tracer sizes at two different temporal scales (10 and 15 min) using the Thermal Infrared (TIR)-1 channel of the INSAT-3DR satellite. The RS-AMVs are able to capture small-scale variations of winds in rapid-scan mode, compared to AMVs retrieved by the operational INSAT-3D satellite available at 30-min intervals. Initial quality assessment of retrieved RS-AMVs is performed using winds from NCEP final analysis, ERA-5 reanalysis, and radiosonde observations. The number of wind vectors are enhanced significantly in RS-AMVs against operational AMVs from INSAT-3DR. An improvement of about 12% in RS-AMVs relative to operational 30-min INSAT-3D-retrieved AMVs has been achieved. Moreover, a sensitivity study has been performed using four-dimensional variational data assimilation experiments to understand the impact of newly retrieved RS-AMVs against operational INSAT-3D-retrieved AMVs for tropical cyclone Fani. Results suggest noteworthy improvements in track prediction when RS-AMVs are used for assimilation as compared to operational AMVs. The model-simulated surface wind structures are also compared with satellite observations for the radius of the maximum winds $$(R_{\max } )$$ and the maximum radial extent of surface winds.

Relationships between Deep Convection Updraft Characteristics and Satellite-Based Super Rapid Scan Mesoscale Atmospheric Motion Vector–Derived Flow

Abstract Rapid acceleration of cloud-top outflow near vigorous storm updrafts can be readily observed in Geostationary Operational Environmental Satellite-14 (GOES-14) super rapid scan (SRS; 60 s) mode data. Conventional wisdom implies that this outflow is related to the intensity of updrafts and the formation of severe weather. However, from an SRS satellite perspective, the pairing of observed expansion and updraft intensity has not been objectively derived and documented. The goal of this study is to relate GOES-14 SRS-derived cloud-top horizontal divergence (CTD) over deep convection to internal updraft characteristics, and document evolution for severe and nonsevere thunderstorms. A new SRS flow derivation system is presented here to estimate storm-scale (<20 km) CTD. This CTD field is coupled with other proxies for storm updraft location and intensity such as overshooting tops (OTs), total lightning flash rates, and three-dimensional flow fields derived from dual-Doppler radar data. Objectively identified OTs with (without) matching CTD maxima were more (less) likely to be associated with radar-observed deep convection and severe weather reports at the ground, suggesting that some OTs were incorrectly identified. The correlation between CTD magnitude, maximum updraft speed, and total lightning was strongly positive for a nonsupercell pulse storm, and weakly positive for a supercell with multiple updraft pulses present. The relationship for the supercell was nonlinear, though larger flash rates are found during periods of larger CTD. Analysis here suggests that combining CTD with OTs and total lightning could have severe weather nowcasting value.

Dynamics of the Interaction of Formic Acid with a Polycrystalline Pt Film Electrode: a Time-Resolved ATR-FTIR Spectroscopy Study at Low Potentials and Temperatures

Aiming at more insights into the interaction of formic acid with a Pt electrode, we have studied the dynamics of formic acid interaction with a polycrystalline Pt film electrode in the potential range around the onset of the reaction, from 0.0 to 0.4 V (reversible hydrogen electrode (RHE)), by a combination of electrochemical and in situ IR spectroscopy transient measurements. The measurements were performed under well-defined mass transport conditions; IR spectra were acquired in an attenuated total reflection (ATR) configuration with a time resolution of up to 25 ms (rapid scan mode). To slow down the reaction kinetics and thus stabilize short-living adsorbed intermediates, measurements were performed at ambient and low reaction temperatures (3 °C). Kinetic H/D isotope effects, introduced by using deuterated formic acid, were explored to learn more about the contribution of C–H bond splitting in the rate-determining step in formic acid dehydration (COad formation). Rapid scan ATR-FTIRS measurements show no measurable time delay between the appearance of the bands related to adsorbed bridge-bonded formate species and adsorbed CO at higher potentials (>0.1 V) and no detectable formate signals at low potentials (≤0.1 V), although COad is still formed even at 0.0 V. Adsorption of HCOOH species at low potentials (0.0–0.1 VRHE) is indicated by a band developing at around 1720 cm−1, which is isotope shifted upon deuteration of the C–H bond. Consequences of these and other observations, including the observation of a bell-shaped potential dependence of the initial rate for COad formation in the potential range 0.0–0.4 V or distinct kinetic H/D isotope effects in the rate constants for COad formation, on the mechanistic understanding of the formic acid–Pt interaction are discussed.

In situ and time-resolved infrared detection of the reactivity induced by electrons in polymer films.

The real time and in situ analysis of chemical reactions such as polymerization reactions, polymer degradation, and oxidation of polymers is of utmost importance. Surprisingly, only few experimental tools allowing this are available. To bridge this gap, we have developed a new experimental setup coupling a 60 keV electron gun with an infrared spectrometer operating in the mid-IR region (800-4000 cm-1) and associated with the rapid scan mode. The measurements are performed using the infrared reflection absorption spectroscopy approach. We worked here with 0.5-25 ms pulses for which the dose per pulse ranges from 4 to 200 kGy. Combining pulse electrons with rapid scan analysis enables performing experiments with a time resolution of 80 ms (and above). Therefore, this new platform can, for instance, give direct insights into reaction mechanisms at stake during the degradation of polymers. First results on a bisphenol A polycarbonate irradiated under vacuum enabling the validation of this new setup and also the recording of time-resolved spectra are presented. Reaction mechanisms are then proposed.

Contrail life cycle and properties from 1 year of MSG/SEVIRI rapid-scan images

The automatic contrail tracking algorithm (ACTA) – developed to automatically follow contrails as they age, drift and spread – enables the study of a large number of contrails and the evolution of contrail properties with time. In this paper we present a year's worth of tracked contrails, from August 2008 to July 2009 in order to derive statistically significant mean values. The tracking is performed using the 5 min rapid-scan mode of the Spinning Enhanced Visible and Infrared Imager (SEVIRI) on board the Meteosat Second Generation (MSG) satellites. The detection is based on the high spatial resolution of the images provided by the Moderate Resolution Imaging Spectroradiometer on board the Terra satellite (Terra/MODIS), where a contrail detection algorithm (CDA) is applied. The results show the satellite-derived average lifetimes of contrails and contrail-cirrus along with the probability density function (PDF) of other geometric characteristics such as mean coverage, distribution and width. In combination with specifically developed algorithms (RRUMS; Rapid Retrieval of Upwelling irradiance from MSG/SEVIRI and COCS (Cirrus Optical properties derived from CALIOP and SEVIRI), explained below) it is possible to derive the radiative forcing (RF), energy forcing (EF), optical thickness (τ) and altitude of the tracked contrails. Mean values here retrieved are duration, 1 h; length, 130 km; width, 8 km; altitude, 11.7 km; optical thickness, 0.34. Radiative forcing and energy forcing are shown for land/water backgrounds in day/night situations.

Influence of Assimilating Satellite-Derived Atmospheric Motion Vector Observations on Numerical Analyses and Forecasts of Tropical Cyclone Track and Intensity

Abstract The influence of assimilating enhanced atmospheric motion vectors (AMVs) on mesoscale analyses and forecasts of tropical cyclones (TC) is investigated. AMVs from the geostationary Multifunctional Transport Satellite (MTSAT) are processed by the Cooperative Institute for Meteorological Satellite Studies (CIMSS, University of Wisconsin–Madison) for the duration of Typhoon Sinlaku (2008), which included a rapid intensification phase and a slow, meandering track. The ensemble Kalman filter and the Weather Research and Forecasting Model are utilized within the Data Assimilation Research Testbed. In addition to conventional observations, three different groups of AMVs are assimilated in parallel experiments: CTL, the same dataset assimilated in the NCEP operational analysis; CIMSS(h), hourly datasets processed by CIMSS; and CIMSS(h+RS), the dataset including AMVs from the rapid-scan mode. With an order of magnitude more AMV data assimilated, the CIMSS(h) analyses exhibit a superior track, intensity, and structure to CTL analyses. The corresponding 3-day ensemble forecasts initialized with CIMSS(h) yield smaller track and intensity errors than those initialized with CTL. During the period when rapid-scan AMVs are available, the CIMSS(h+RS) analyses offer additional modifications to the TC and its environment. In contrast to many members in the ensemble forecasts initialized from the CTL and CIMSS(h) analyses that predict an erroneous landfall in China, the CIMSS(h+RS) members capture recurvature, albeit prematurely. The results demonstrate the promise of assimilating enhanced AMV data into regional TC models. Further studies to identify optimal strategies for assimilating integrated full-resolution multivariate data from satellites are under way.

A fast method for the retrieval of integrated longwave and shortwave top-of-atmosphere upwelling irradiances from MSG/SEVIRI (RRUMS)

Abstract. A new Rapid Retrieval of Upwelling irradiances from MSG/SEVIRI (RRUMS) is presented. It has been developed to observe the top-of-atmosphere irradiances of small scale and rapidly changing features that are not sufficiently resolved by specific Earth radiation budget sensors. Our retrieval takes advantage of the spatial and temporal resolution of MSG/SEVIRI and provides outgoing longwave and reflected shortwave radiation only by means of a combination of SEVIRI channels. The longwave retrieval is based on a simple linear combination of brightness temperatures from the SEVIRI infrared channels. The shortwave retrieval is based on a neural network that requires as input the visible and near-infrared SEVIRI channels. Both LW and SW algorithms have been validated by comparing their results with CERES and GERB irradiance observations. While being less accurate than their dedicated counterparts, the SEVIRI-based methods have two major advantages compared to CERES and GERB: their higher spatial resolution and the better temporal resolution. With our retrievals it is possible to observe the radiative effect of small-scale features such as cumulus clouds, cirrus clouds, or aircraft contrails. The spatial resolution of SEVIRI is 3 km × 3 km in the sub-satellite point, remarkably better than that of CERES (20 km) or GERB (45 km). The temporal resolution is 15 min (5 min in the Rapid-Scan mode), the same as GERB, but significantly better than that of CERES which, being on board of a polar orbiting satellite, has a temporal resolution as low as 2 overpasses per day.

Detection of convective initiation using Meteosat SEVIRI: implementation in and verification with the tracking and nowcasting algorithm Cb-TRAM

Abstract. In this paper a new detection scheme for convective initiation (CI) under day and night conditions is presented. The new algorithm combines the strengths of two existing methods for detecting CI with geostationary satellite data. It uses the channels of the Spinning Enhanced Visible and Infrared Imager (SEVIRI) onboard Meteosat Second Generation (MSG). For the new algorithm five infrared (IR) criteria from the Satellite Convection Analysis and Tracking algorithm (SATCAST) and one high-resolution visible channel (HRV) criteria from Cb-TRAM were adapted. This set of criteria aims to identify the typical development of quickly developing convective cells in an early stage. The different criteria include time trends of the 10.8 IR channel, and IR channel differences, as well as their time trends. To provide the trend fields an optical-flow-based method is used: the pyramidal matching algorithm, which is part of Cb-TRAM. The new detection scheme is implemented in Cb-TRAM, and is verified for seven days which comprise different weather situations in central Europe. Contrasted with the original early-stage detection scheme of Cb-TRAM, skill scores are provided. From the comparison against detections of later thunderstorm stages, which are also provided by Cb-TRAM, a decrease in false prior warnings (false alarm ratio) from 91 to 81% is presented, an increase of the critical success index from 7.4 to 12.7%, and a decrease of the BIAS from 320 to 146% for normal scan mode. Similar trends are found for rapid scan mode. Most obvious is the decline of false alarms found for the synoptic class "cold air" masses.

Light-dark Adaptation of Channelrhodopsin C128T Mutant

Channelrhodopsins are microbial type rhodopsins that operate as light-gated ion channels. Largely prolonged lifetimes of the conducting state of channelrhodopsin-2 may be achieved by mutations of crucial single amino acids, i.e. cysteine 128. Such mutants are of great scientific interest in the field of neurophysiology because they allow neurons to be switched on and off on demand (step function rhodopsins). Due to their slow photocycle, structural alterations of these proteins can be studied by vibrational spectroscopy in more detail than possible with wild type. Here, we present spectroscopic evidence that the photocycle of the C128T mutant involves three different dark-adapted states that are populated according to the wavelength and duration of the preceding illumination. Our results suggest an important role of multiphoton reactions and the previously described side reaction for dark state regeneration. Structural changes that cause formation and depletion of the assumed ion conducting state P520 are only small and follow larger changes that occur early and late in the photocycle, respectively. They require only minor structural rearrangements of amino acids near the retinal binding pocket and are triggered by all-trans/13-cis retinal isomerization, although additional isomerizations are also involved in the photocycle. We will discuss an extended photocycle model of this mutant on the basis of spectroscopic and electrophysiological data.

Comparison between GOES-12 Overshooting-Top Detections, WSR-88D Radar Reflectivity, and Severe Storm Reports

Abstract Studies have found that convective storms with overshooting-top (OT) signatures in weather satellite imagery are often associated with hazardous weather, such as heavy rainfall, tornadoes, damaging winds, and large hail. An objective satellite-based OT detection product has been developed using 11-μm infrared window (IRW) channel brightness temperatures (BTs) for the upcoming R series of the Geostationary Operational Environmental Satellite (GOES-R) Advanced Baseline Imager. In this study, this method is applied to GOES-12 IRW data and the OT detections are compared with radar data, severe storm reports, and severe weather warnings over the eastern United States. The goals of this study are to 1) improve forecaster understanding of satellite OT signatures relative to commonly available radar products, 2) assess OT detection product accuracy, and 3) evaluate the utility of an OT detection product for diagnosing hazardous convective storms. The coevolution of radar-derived products and satellite OT signatures indicates that an OT often corresponds with the highest radar echo top and reflectivity maximum aloft. Validation of OT detections relative to composite reflectivity indicates an algorithm false-alarm ratio of 16%, with OTs within the coldest IRW BT range (<200 K) being the most accurate. A significant IRW BT minimum typically present with an OT is more often associated with heavy precipitation than a region with a spatially uniform BT. Severe weather was often associated with OT detections during the warm season (April–September) and over the southern United States. The severe weather to OT relationship increased by 15% when GOES operated in rapid-scan mode, showing the importance of high temporal resolution for observing and detecting rapidly evolving cloud-top features. Comparison of the earliest OT detection associated with a severe weather report showed that 75% of the cases occur before severe weather and that 42% of collocated severe weather reports had either an OT detected before a severe weather warning or no warning issued at all. The relationships between satellite OT signatures, severe weather, and heavy rainfall shown in this paper suggest that 1) when an OT is detected, the particular storm is likely producing heavy rainfall and/or possibly severe weather; 2) an objective OT detection product can be used to increase situational awareness and forecaster confidence that a given storm is severe; and 3) this product may be particularly useful in regions with insufficient radar coverage.

Fourier transform infrared spectroscopy approach for measurements of photoluminescence and electroluminescence in mid-infrared

An improved Fourier transform infrared spectroscopy approach adapting to photoluminescence and electroluminescence measurements in mid-infrared has been developed, in which diode-pumped solid-state excitation lasers were adopted for photoluminescence excitation. In this approach, three different Fourier transform infrared modes of rapid scan, double modulation, and step scan were software switchable without changing the hardware or connections. The advantages and limitations of each mode were analyzed in detail. Using this approach a group of III-V and II-VI samples from near-infrared extending to mid-infrared with photoluminescence intensities in a wider range have been characterized at room temperature to demonstrate the validity and overall performances of the system. The weaker electroluminescence of quantum cascade lasers in mid-infrared band was also surveyed at different resolutions. Results show that for samples with relatively strong photoluminescence or electroluminescence out off the background, rapid scan mode is the most preferable. For weaker photoluminescence or electroluminescence overlapped with background, double modulation is the most effective mode. To get a better signal noise ratio when weaker photoluminescence or electroluminescence signal has been observed in double modulation mode, switching to step scan mode should be an advisable option despite the long data acquiring time and limited resolution.

The accuracy of dynamic predictive autofocusing for whole slide imaging

Context: Whole slide imaging (WSI) for digital pathology involves the rapid automated acquisition of multiple high-power fields from a microscope slide containing a tissue specimen. Capturing each field in the correct focal plane is essential to create high-quality digital images. Others have described a novel focusing method which reduces the number of focal planes required to generate accurate focus. However, this method was not applied dynamically in an automated WSI system under continuous motion. Aims: This report measures the accuracy of this method when applied in a rapid continuous scan mode using a dual sensor WSI system with interleaved acquisition of images. Methods: We acquired over 400 tiles in a “stop and go” scan mode, surveying the entire z depth in each tile and used this as ground truth. We compared this ground truth focal height to the focal height determined using a rapid 3-point focus algorithm applied dynamically in a continuous scanning mode. Results: Our data showed the average focal height error of 0.30 (±0.27) μm compared to ground truth, which is well within the system’s depth of field. On a tile by tile assessment, approximately 95% of the tiles were within the system’s depth of field. Further, this method was six times faster than acquiring tiles compared to the same method in a non-continuous scan mode. Conclusions: The data indicates that the method employed can yield a significant improvement in scan speed while maintaining highly accurate autofocusing.

An automatic contrail tracking algorithm

Abstract. A method designed to track the life cycle of contrail-cirrus using satellite data with high temporal and spatial resolution, from its formation to the final dissolution of the aviation-induced cirrus cloud is presented. The method follows the evolution of contrails from their linear stage until they are undistinguishable from natural cirrus clouds. Therefore, the study of the effect of aircraft-induced clouds in the atmosphere is no longer restricted to linear contrails and can include contrail-cirrus. The method takes advantage of the high spatial resolution of polar orbiting satellites and the high temporal resolution of geostationary satellites to identify the pixels that belong to an aviation induced cloud. The high spatial resolution data of the MODIS sensor is used for contrail detection, and the high temporal resolution of the SEVIRI sensor in the Rapid Scan mode is used for contrail tracking. An example is included in which the method is applied to the study of a long lived contrail over the bay of Biscay.

Use of a Vertical Vorticity Equation in Variational Dual-Doppler Wind Analysis

Abstract The utility of the anelastic vertical vorticity equation in a weak-constraint (least squares error) variational dual-Doppler wind analysis procedure is explored. The analysis winds are obtained by minimizing a cost function accounting for the discrepancies between observed and analyzed radial winds, errors in the mass conservation equation, errors in the anelastic vertical vorticity equation, and spatial smoothness constraints. By using Taylor’s frozen-turbulence hypothesis to shift analysis winds to observation points, discrepancies between radially projected analysis winds and radial wind observations can be calculated at the actual times and locations the data are acquired. The frozen-turbulence hypothesis is also used to evaluate the local derivative term in the vorticity equation. Tests of the analysis procedure are performed with analytical pseudo-observations of an array of translating and temporally decaying counterrotating updrafts and downdrafts generated from a Beltrami flow solution of the Navier–Stokes equations. The experiments explore the value added to the analysis by the vorticity equation constraint in the common scenario of substantial missing low-level data (radial wind observations at heights beneath 1.5 km are withheld from the analysis). Experiments focus on the sensitivity of the most sensitive analysis variable—the vertical velocity component—to values of the weighting coefficients, volume scan period, number of volume scans, and errors in the estimated frozen-turbulence pattern-translation components. Although the vorticity equation constraint is found to add value to many of these analyses, the analysis can become significantly degraded if estimates of the pattern-translation components are largely in error or if the frozen-turbulence hypothesis itself breaks down. However, tests also suggest that these negative impacts can be mitigated if data are available in a rapid-scan mode.

Detection of a gem-diamine and a stable quinonoid intermediate in the reaction catalyzed by serine–glyoxylate aminotransferase from Hyphomicrobium methylovorum

BackgroundThe enzyme l-serine–glyoxylate aminotransferase (SGAT) from Hyphomicrobium methylovorum is a PLP-containing enzyme that catalyzes the conversion of l-serine and glyoxylate to hydroxypyruvate and glycine. The cloned enzyme expressed in Escherichia coli is isolated as a mixture of the E:PLP and E:PMP forms. The PLP form of the enzyme has a maximum absorbance at 413 nm. MethodsUv–visible spectra of SGAT were obtained using an HP-8453 diode array spectrophotometer in the absence and presence of substrates and substrate analogs. Pre-steady state kinetic studies were carried out using an OLIS rapid scanning spectrophotometer in the rapid scanning mode. ResultsIncubation of the enzyme with a saturating concentration D-serine leads to a shift in the 413 nm peak to 421 nm that is ascribed to the external aldimine. The reverse stereochemistry of D-serine does not allow for abstraction of the Cα proton by the ε-amine of the active site lysine residue leading to an abortive external aldimine intermediate. Pre-steady state studies pushing SGAT against d-serine leads to a rapid decrease in the 413 nm peak and an increase at ∼330 nm with an associated rate constant of 47 s−1 at pH 7.6. This is followed by a slower decrease (0.26 s−1) at 330 nm and an increase and shift of the 413 nm peak to 421 nm. The intermediate species that absorbs at ∼330 nm is attributed to the gem-diamine intermediate. The rate of the fast phase increases with pH and increase in rate is likely due to the deprotonation of an enzymatic group that accepts a proton from the α-amine of d-serine. In the presence of hydroxypyruvate and ammonia the enzyme spectra display an increase in absorbance at 521 nm that occurs on the order of minutes. The shape and position of the 521 nm species is consistent with a quinonoid intermediate. General significanceThe data suggest a non-enzymatic reaction between hydroxypyruvate and ammonia to form an imine which will be in equilibrium with the enamine. A mechanism is proposed by which the enamine reacts with the PLP form of SGAT to generate the stable highly conjugated quinonoid intermediate.

Monitoring Light-induced Structural Changes of Channelrhodopsin-2 by UV-visible and Fourier Transform Infrared Spectroscopy

Channelrhodopsin-2 (ChR2) is a microbial type rhodopsin and a light-gated cation channel that controls phototaxis in Chlamydomonas. We expressed ChR2 in COS-cells, purified it, and subsequently investigated this unusual photoreceptor by flash photolysis and UV-visible and Fourier transform infrared difference spectroscopy. Several transient photoproducts of the wild type ChR2 were identified, and their kinetics and molecular properties were compared with those of the ChR2 mutant E90Q. Based on the spectroscopic data we developed a model of the photocycle comprising six distinguishable intermediates. This photocycle shows similarities to the photocycle of the ChR2-related Channelrhodopsin of Volvox but also displays significant differences. We show that molecular changes include retinal isomerization, changes in hydrogen bonding of carboxylic acids, and large alterations of the protein backbone structure. These alterations are stronger than those observed in the photocycle of other microbial rhodopsins like bacteriorhodopsin and are related to those occurring in animal rhodopsins. UV-visible and Fourier transform infrared difference spectroscopy revealed two late intermediates with different time constants of tau = 6 and 40 s that exist during the recovery of the dark state. The carboxylic side chain of Glu(90) is involved in the slow transition. The molecular changes during the ChR2 photocycle are discussed with respect to other members of the rhodopsin family.