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Functional derivatives applied to error propagation of uncertainties in topography to large-aperture scintillometer-derived heat fluxes
Gruber, M.A. ; Fochesatto, G.J. ; Hartogensis, O.K. ; Lysy, M. - \ 2014
Atmospheric Measurement Techniques 7 (2014). - ISSN 1867-1381 - p. 2361 - 2371.
structure parameter - refractive-index - effective height - surface
Scintillometer measurements allow for estimations of the refractive index structure parameter Cn2 over large areas in the atmospheric surface layer. Turbulent fluxes of heat and momentum are inferred through coupled sets of equations derived from the Monin–Obukhov similarity hypothesis. One-dimensional sensitivity functions have been produced that relate the sensitivity of heat fluxes to uncertainties in single values of beam height over flat terrain. However, real field sites include variable topography. We develop here, using functional derivatives, the first analysis of the sensitivity of scintillometer-derived sensible heat fluxes to uncertainties in spatially distributed topographic measurements. Sensitivity is shown to be concentrated in areas near the center of the beam path and where the underlying topography is closest to the beam height. Relative uncertainty contributions to the sensible heat flux from uncertainties in topography can reach 20% of the heat flux in some cases. Uncertainty may be greatly reduced by focusing accurate topographic measurements in these specific areas. A new two-dimensional variable terrain sensitivity function is developed for quantitative error analysis. This function is compared with the previous one-dimensional sensitivity function for the same measurement strategy over flat terrain. Additionally, a new method of solution to the set of coupled equations is produced that eliminates computational error.
Rise-Time of FRET-Acceptor Fluorescence Tracks Protein Folding
Lindhoud, S. ; Westphal, A.H. ; Mierlo, C.P.M. van; Visser, A.J.W.G. ; Borst, J.W. - \ 2014
International Journal of Molecular Sciences 15 (2014)12. - ISSN 1661-6596 - p. 23836 - 23850.
resonance energy-transfer - beta parallel protein - single-molecule fluorescence - azotobacter-vinelandii - spectroscopic ruler - refractive-index - tryptophan residue - molten globule - wild-type - pathway
Uniform labeling of proteins with fluorescent donor and acceptor dyes with an equimolar ratio is paramount for accurate determination of Förster resonance energy transfer (FRET) efficiencies. In practice, however, the labeled protein population contains donor-labeled molecules that have no corresponding acceptor. These FRET-inactive donors contaminate the donor fluorescence signal, which leads to underestimation of FRET efficiencies in conventional fluorescence intensity and lifetime-based FRET experiments. Such contamination is avoided if FRET efficiencies are extracted from the rise time of acceptor fluorescence upon donor excitation. The reciprocal value of the rise time of acceptor fluorescence is equal to the decay rate of the FRET-active donor fluorescence. Here, we have determined rise times of sensitized acceptor fluorescence to study the folding of double-labeled apoflavodoxin molecules and show that this approach tracks the characteristics of apoflavodoxin's complex folding pathway.
The Effect of a New Calibration Procedure on the Measurement Accuracy of Scintec's Displaced-Beam Laser Scintillometer
Kesteren, A.J.H. van; Hartogensis, O.K. ; Kroonenberg, A.C. van den - \ 2014
Boundary-Layer Meteorology 151 (2014)2. - ISSN 0006-8314 - p. 257 - 271.
atmospheric surface-layer - stable boundary-layer - optical scintillation - inner scale - structure parameter - flux measurements - refractive-index - sensible heat - turbulence - spectrum
We describe a new calibration procedure included in the production process of Scintec’s displaced-beam laser scintillometers (SLS-20/40) and its effect on their measurement accuracy. The calibration procedure determines the factual displacement distances of the laser beams at the receiver and transmitter units, instead of assuming a prescribed displacement distance of 2.70 mm. For this study, four scintillometers operated by Wageningen University and the German Meteorological Service were calibrated by Scintec and their data re-analyzed. The results show that significant discrepancies may exist between the factual and the prescribed displacement distances. Generally, the factual displacement is about 0.1 mm smaller than 2.70 mm, but extremes varied between 0.04 and 0.24 mm. Correspondingly, using non-calibrated scintillometers may result in biases as large as 20 % in the estimates of the inner-scale length, l0, the structure parameter of the refractive index, Cn2, and the friction velocity, u*. The bias in the sensible heat flux was negligible, because biases in Cn2 and u* cancel. Hence, the discrepancies explain much of the long observed underestimations of u * determined by these scintillometers. Furthermore, the calibration improves the mutual agreement between the scintillometers for l 0 , but especially for Cn2. Finally, it is noted that the measurement specifications of the scintillometer do not expire and hence the results of the calibration can be applied retroactively
Derivation of Structure Parameters of Temperature and Humidity in the Convective Boundary Layer from Large-Eddy Simulations and Implications for the Interpretation of Scintillometer Observations
Maronga, B. ; Moene, A.F. ; Dinther, D. van; Raasch, S. ; Bosveld, F.C. ; Gioli, B. - \ 2013
Boundary-Layer Meteorology 148 (2013)1. - ISSN 0006-8314 - p. 1 - 30.
large-aperture scintillometer - heterogeneous land-surface - flevoland field experiment - index-structure parameter - refractive-index - optical scintillometer - water-vapor - fluxes - model - heat
We derive the turbulent structure parameters of temperature C 2 T and humidity C 2 q from high-resolution large-eddy simulations (LES) of a homogeneously-heated convective boundary layer. Boundary conditions and model forcing were derived from measurements at Cabauw in The Netherlands. Three different methods to obtain the structure-parameters from LES are investigated. The shape of the vertical structure-parameter profiles from all three methods compare well with former experimental and LES results. Depending on the method, deviations in the magnitude up to a factor of two are found and traced back to the effects of discretization and numerical dissipation of the advection scheme. Furthermore, we validate the LES data with airborne and large-aperture scintillometer (LAS) measurements at Cabauw. Virtual path measurements are used to study the variability of C 2 T in the mixed layer and surface layer and its implications for airborne and LAS measurements. A high variability of C 2 T along a given horizontal path in the LES data is associated with plumes (high values) and downdrafts (low values). The path average of C 2 T varies rapidly in time due to the limited path length. The LES results suggest that measured path averages require sufficient temporal averaging and an adequate ratio of path length to height above the ground for the LAS in order to approach the domain average of C 2 T .
A critical revision of the estimation of the latent heat flux from two-wavelength scintillometry
Ward, H.C. ; Evans, J.G. ; Hartogensis, O.K. ; Moene, A.F. ; Debruin, H.A.R. ; Grimmond, C.S.B. - \ 2013
Quarterly Journal of the Royal Meteorological Society 139 (2013)676. - ISSN 0035-9009 - p. 1912 - 1922.
large-aperture scintillometer - heterogeneous land-surface - temperature-humidity correlation - refractive-index - water-vapor - structure parameters - regional advection - eddy-covariance - scintillation - fluctuations
Simultaneous scintillometer measurements at multiple wavelengths (pairing visible or infrared with millimetre or radio waves) have the potential to provide estimates of path-averaged surface fluxes of sensible and latent heat. Traditionally, the equations to deduce fluxes from measurements of the refractive index structure parameter at the two wavelengths have been formulated in terms of absolute humidity. Here, it is shown that formulation in terms of specific humidity has several advantages. Specific humidity satisfies the requirement for a conserved variable in similarity theory and inherently accounts for density effects misapportioned through the use of absolute humidity. The validity and interpretation of both formulations are assessed and the analogy with open-path infrared gas analyser density corrections is discussed. Original derivations using absolute humidity to represent the influence of water vapour are shown to misrepresent the latent heat flux. The errors in the flux, which depend on the Bowen ratio (larger for drier conditions), may be of the order of 10%. The sensible heat flux is shown to remain unchanged. It is also verified that use of a single scintillometer at optical wavelengths is essentially unaffected by these new formulations. Where it may not be possible to reprocess two-wavelength results, a density correction to the latent heat flux is proposed for scintillometry, which can be applied retrospectively to reduce the error.
Illuminating the off-pathway nature of the molten globule folding intermediate of an a-ß parallel protein
Lindhoud, S. ; Westphal, A.H. ; Borst, J.W. ; Mierlo, C.P.M. van - \ 2012
PLoS ONE 7 (2012)9. - ISSN 1932-6203
azotobacter-vinelandii apoflavodoxin - refractive-index - fluorescence depolarization - spectroscopic ruler - hydrogen-exchange - energy landscape - state - flavodoxin - aggregation - mechanism
Partially folded protein species transiently form during folding of most proteins. Often, these species are molten globules, which may be on- or off-pathway to the native state. Molten globules are ensembles of interconverting protein conformers that have a substantial amount of secondary structure, but lack virtually all tertiary side-chain packing characteristics of natively folded proteins. Due to solvent-exposed hydrophobic groups, molten globules are prone to aggregation, which can have detrimental effects on organisms. The molten globule observed during folding of the 179-residue apoflavodoxin from Azotobacter vinelandii is off-pathway, as it has to unfold before native protein can form. Here, we study folding of apoflavodoxin and characterize its molten globule using fluorescence spectroscopy and Förster Resonance Energy Transfer (FRET). Apoflavodoxin is site-specifically labeled with fluorescent donor and acceptor dyes, utilizing dye-inaccessibility of Cys69 in cofactor-bound protein. Donor (i.e., Alexa Fluor 488) is covalently attached to Cys69 in all apoflavodoxin variants used. Acceptor (i.e., Alexa Fluor 568) is coupled to Cys1, Cys131 and Cys178, respectively. Our FRET data show that apoflavodoxin’s molten globule forms in a non-cooperative manner and that its N-terminal 69 residues fold last. In addition, striking conformational differences between molten globule and native protein are revealed, because the inter-label distances sampled in the 111-residue C-terminal segment of the molten globule are shorter than observed for native apoflavodoxin. Thus, FRET sheds light on the off-pathway nature of the molten globule during folding of an a-ß parallel protein
On Monin–Obukhov Scaling in and Above the Atmospheric Surface Layer: The Complexities of Elevated Scintillometer Measurements
Braam, M. ; Bosveld, F.C. ; Moene, A.F. - \ 2012
Boundary-Layer Meteorology 144 (2012)2. - ISSN 0006-8314 - p. 157 - 177.
large-aperture scintillometer - boundary-layer - structure parameters - sonic anemometer - refractive-index - sensible heat - water-vapor - temperature - fluxes - humidity
In scintillometry Monin–Obukhov similarity theory (MOST) is used to calculate the surface sensible heat flux from the structure parameter of temperature (CT2)(CT2) . In order to prevent saturation a scintillometer can be installed at an elevated level. However, in that case the observation level might be located outside the atmospheric surface layer (ASL) and thus the validity of MOST questioned. Therefore, we examine two concepts to determine the turbulent surface sensible heat flux from the structure parameter at elevated levels with data obtained at 60-m height on the Cabauw tower (the Netherlands). In the first concept (MOSTs) CT2CT2 is still scaled with the surface flux, whereas in the second (MOSTl) CT2CT2 is scaled with the local sensible heat flux. The CT2CT2 obtained from both concepts is compared with direct observations of CT2CT2 using a sonic anemometer/thermometer. In the afternoon (when the measurement height is located within the ASL) both concepts give results that are comparable to the directly observed values of CT2CT2 . In the morning (data outside the ASL), our data do not unequivocally support either of the two concepts. First, the peak in CT2CT2 that occurs when the measurement height is located in the entrainment zone disqualifies the use of MOST. Second, during the morning transition, local scaling shows the correct pattern (zero flux and a minimum in CT2CT2) but underestimates CT2CT2 by a factor of ten. Third, from the best linear fit a we found that the slope of MOSTl gave better results, whereas the offset is closer to zero for MOSTs. Further, the correlation between the direct observations and MOST-scaled results is low and similar for the two concepts. In the end, we conclude that MOST is not applicable for the morning hours when the observation level is above the ASL.
Green-Fluorescent Protein from the Bioluminescent Jellyfish Clytia gregaria Is an Obligate Dimer and Does Not Form a Stable Complex with the Ca2+-Discharged Photoprotein Clytin.
Malikova, N.P. ; Visser, N.V. ; Hoek, A. van; Skakun, V.V. ; Vysotski, E.S. ; Lee, J. ; Visser, A.J.W.G. - \ 2011
Biochemistry 50 (2011)20. - ISSN 0006-2960 - p. 4232 - 4241.
vibrio-fischeri y1 - energy-transfer - correlation spectroscopy - bacterial luciferase - refractive-index - photobacterium-leiognathi - polarized fluorescence - excitation transfer - recombinant obelin - lumazine protein
Green-fluorescent protein (GFP) is the origin of the green bioluminescence color exhibited by several marine hydrozoans and anthozoans. The mechanism is believed to be Fo¨rster resonance energy transfer (FRET) within a luciferase-GFP or photoprotein-GFP complex. As the effect is found in vitro at micromolar concentrations, for FRET to occur this complex must have an affinity in the micromolar range. We present here a fluorescence dynamics investigation of the recombinant bioluminescence proteins from the jellyfish Clytia gregaria, the photoprotein clytin in its Ca2+-discharged form that is highly fluorescent (¿max = 506 nm) and its GFP (cgreGFP; ¿max = 500 nm). Ca2+-discharged clytin shows a predominant fluorescence lifetime of 5.7 ns, which is assigned to the final emitting state of the bioluminescence reaction product, coelenteramide anion, and a fluorescence anisotropy decay or rotational correlation time of 12 ns (20 °C), consistent with tight binding and rotation with the whole protein. A 34 ns correlation time combined with a translational diffusion constant and molecular brightness from fluorescence fluctuation spectroscopy all confirm that cgreGFP is an obligate dimer down to nanomolar concentrations. Within the dimer, the two chromophores have a coupled excited-state transition yielding fluorescence depolarization via FRET with a transfer correlation time of 0.5 ns. The 34 ns time of cgreGFP showed no change upon addition of a 1000-fold excess of Ca2+-discharged clytin, indicating no stable complexation below 0.2 mM. It is proposed that any bioluminescence FRET complex with micromolar affinity must be one formed transiently by the cgreGFP dimer with a short-lived (millisecond) intermediate in the clytin reaction pathway.
Analysis of the Systematic Errors Found in the Kipp & Zonen Large-Aperture Scintillometer
Kesteren, A.J.H. van; Hartogensis, O.K. - \ 2011
Boundary-Layer Meteorology 138 (2011)3. - ISSN 0006-8314 - p. 493 - 509.
heterogeneous land-surface - sensible heat fluxes - optical scintillometer - refractive-index - c-n(2) - area
Studies have shown a systematic error in the Kipp & Zonen large-aperture scintillometer (K&ZLAS) measurements of the sensible heat flux, H. We improved on these studies and compared four K&ZLASs with a Wageningen large-aperture scintillometer at the Chilbolton Observatory. The scintillometers were installed such that their footprints were the same and independent flux measurements were made along the measurement path. This allowed us to compare H and the direct scintillometer output, the refractive index structure parameter, C2n. Furthermore, spectral analysis was performed on the raw scintillometer signal to investigate the characteristics of the error. Firstly, correlation coefficients =0.99 confirm the robustness of the scintillometer method, and secondly we discovered two systematic errors: the low-C2n error and the high-C2n error. The low-C2n error is a non-linear error that is caused by high-frequency noise, and we suspect the error to be caused by the calibration circuit in the receiver. It varies between each K&ZLAS, is significant for H = 50 W m-2, and we propose a solution to remove this error using the demodulated signal. The high-C2n error identified by us is the systematic error found in previous studies.We suspect this error to be caused by poor focal alignment of the receiver detector and the transmitter light-emitting diode that causes ineffective use of the Fresnel lens in the current Kipp & Zonen design. It varies between each K&ZLAS (35% up to 240%) and can only be removed by comparing with a reference scintillometer in the field.
Global analysis of Förster resonance energy transfer in live cells measured by fluorescence lifetime imaging microscopy exploiting the rise time of acceptor fluorescence
Laptenok, S. ; Borst, J.W. ; Mullen, K.M. ; Stokkum, I.H.M. van; Visser, A.J.W.G. ; Amerongen, H. van - \ 2010
Physical Chemistry Chemical Physics 12 (2010)27. - ISSN 1463-9076 - p. 7593 - 7602.
nonlinear least-squares - quantitative fret analysis - resolved spectra - correlation spectroscopy - picosecond fluorescence - reference convolution - refractive-index - proteins - excitation - precision
A methodology is described for the quantitative determination of Förster resonance energy transfer (FRET) in live cells using the rise time of acceptor fluorescence as determined with fluorescence lifetime imaging microscopy (FLIM). An advantage of this method is that only those molecules that are involved in the energy-transfer process are monitored. This contrasts with current methods that measure either steady-state fluorescence of donor and acceptor molecules or time-resolved fluorescence of donor molecules, and thereby probe a mixture of donor molecules that are involved in FRET and those that are fluorescent but not involved in FRET. The absence of FRET can, for instance, be due to unwanted acceptor bleaching or incomplete maturing of visible proteins that should act as acceptor molecules. In addition, parameters describing the rise of acceptor fluorescence and the decay of donor fluorescence can be determined via simultaneous global analysis of multiple FLIM images, thereby increasing the reliability of the analysis. In the present study, plant protoplasts transfected with fusions of visible fluorescent proteins are used to illustrate the new data analysis method. It is demonstrated that the distances estimated with the present method are substantially smaller than those estimated from the average donor lifetimes, due to a fraction of non-transferring donor molecules. Software to reproduce the presented results is provided in an open-source and freely available package called "TIMP" for "The R project for Statistical Computing"
Structural properties and gelatinisation characteristics of potato and cassava starches and mutants thereof
Gomand, S.V. ; Lamberts, L. ; Derde, L.J. ; Groesaert, H. ; Vandeputte, G.E. ; Goderis, B. ; Visser, R.G.F. ; Delcour, J.A. - \ 2010
Food Hydrocolloids 24 (2010)4. - ISSN 0268-005X - p. 307 - 317.
aspects provide insight - size-exclusion chromatography - amylose maize starches - molecular-structure - rice starches - retrogradation properties - amylopectin structure - light-scattering - refractive-index - granular starch
The molecular size of amylopectin (AP) and amylose (AM), AP chain length distribution, crystallinity and granular structure (morphology and granule size distribution) of five wild type potato starches (wtps), five AM free potato starches (amfps), four high-AM potato starches (haps), one wild type cassava starch (wtcs) and one AM free cassava starch (amfcs) were investigated and related to their gelatinisation characteristics. Starches with higher levels of short chains [degree of polymerisation (DP) 6–9 and DP 10–14)] had lower gelatinisation onset (To), peak (Tp) and conclusion (Tc) temperatures, whereas higher contents of longer chains (DP 18–25 and DP 25–80) led to higher gelatinisation temperatures. Gelatinisation enthalpies (¿H) increased with degree of crystallinity. The granules of wtps were larger than those of amfps and haps, respectively. No differences in morphology were observed between wtps and amfps granules, but the haps granules had more irregular surfaces and showed multi-lobed granules.
Exploring the structure of the N-terminal domain of CP29 with ultrafast fluorescence spectroscopy
Berghuis, B.A. ; Spruijt, R.B. ; Koehorst, R.B.M. ; Hoek, A. van; Laptenok, S. ; Oort, B.F. van; Amerongen, H. van - \ 2010
European Biophysics Journal 39 (2010)4. - ISSN 0175-7571 - p. 631 - 638.
light-harvesting complex - radical-cation formation - plant antenna protein - energy-transfer - photosystem-ii - green plants - escherichia-coli - refractive-index - chlorophyll-a - pump-probe
A high-throughput Förster resonance energy transfer (FRET) study was performed on the approximately 100 amino acids long N-terminal domain of the photosynthetic complex CP29 of higher plants. For this purpose, CP29 was singly mutated along its N-terminal domain, replacing one-by-one native amino acids by a cysteine, which was labeled with a BODIPY fluorescent probe, and reconstituted with the natural pigments of CP9, chlorophylls and xanthophylls. Picosecond fluorescence experiments revealed rapid energy transfer (~20–70 ps) from BODIPY at amino-acid positions 4, 22, 33, 40, 56, 65, 74, 90, and 97 to Chl a molecules in the hydrophobic part of the protein. From the energy transfer times, distances were estimated between label and chlorophyll molecules, using the Förster equation. When the label was attached to amino acids 4, 56, and 97, it was found to be located very close to the protein core (~15 Å), whereas labels at positions 15, 22, 33, 40, 65, 74, and 90 were found at somewhat larger distances. It is concluded that the entire N-terminal domain is in close contact with the hydrophobic core and that there is no loop sticking out into the stroma. Most of the results support a recently proposed topological model for the N-terminus of CP29, which was based on electron-spin-resonance measurements on spin-labeled CP29 with and without its natural pigment content. The present results lead to a slight refinement of that model
Ultrafast resonance energy transfer from a site-specifically attached fluorescent chromophore reveals the folding of the N-terminal domain of CP29
Oort, B.F. van; Murali, S. ; Wientjes, E. ; Koehorst, R.B.M. ; Spruijt, R.B. ; Hoek, A. van; Croce, R. ; Amerongen, H. van - \ 2009
Chemical Physics 357 (2009)1-3. - ISSN 0301-0104 - p. 113 - 119.
light-harvesting complexes - photosystem-ii subunit - chlorophyll a/b complex - plant antenna protein - green plants - absorption properties - escherichia-coli - refractive-index - excited-states - pump-probe
The photosynthetic minor antenna complex CP29 of higher plants was singly mutated, overexpressed in Escherichia coli, selectively labeled with the fluorescent dye TAMRA at three positions in the N-terminal domain, and reconstituted with its natural pigments. Picosecond fluorescence experiments revealed rapid excitation energy transfer (20 ps) from TAMRA covalently attached to a cysteine at either position 4 or 97 (near the beginning and end of the N-terminal domain) to the chlorophylls in the hydrophobic part of the protein. This indicates that the N-terminus is folded back on the hydrophobic core. In 20% of the complexes, efficient transfer was lacking, indicating that the N-terminus can adopt different conformations. Time-resolved polarized fluorescence measurements demonstrate that the non-transferring conformations only allow restricted rotational motion of the dye molecule. When TAMRA was attached to a cysteine at position 40, the overall transfer efficiency was far lower, reflecting a larger distance to the hydrophobic region
Macromolecular crowding compacts unfolded apoflavodoxin and causes severe aggregation of the off-pathway intermediate during apoflavodoxin folding
Engel, R. ; Westphal, A.H. ; Huberts, D. ; Nabuurs, S.M. ; Lindhoud, S. ; Visser, A.J.W.G. ; Mierlo, C.P.M. van - \ 2008
Journal of Biological Chemistry 283 (2008)41. - ISSN 0021-9258 - p. 27383 - 27394.
fluorescence correlation spectroscopy - azotobacter-vinelandii apoflavodoxin - inclusion-body formation - protein stability - hydrogen-exchange - escherichia-coli - molecular chaperones - refractive-index - self-association - creatine-kinase
To understand how proteins fold in vivo, it is important to investigate the effects of macromolecular crowding on protein folding. Here, the influence of crowding on in vitro apoflavodoxin folding, which involves a relatively stable off-pathway intermediate with molten globule characteristics, is reported. To mimic crowded conditions in cells, dextran 20 at 30% (w/v) is used, and its effects are measured by a diverse combination of optical spectroscopic techniques. Fluorescence correlation spectroscopy shows that unfolded apoflavodoxin has a hydrodynamic radius of 37 +/- 3 angstrom at 3M guanidine hydrochloride. Forster resonance energy transfer measurements reveal that subsequent addition of dextran 20 leads to a decrease in protein volume of about 29%, which corresponds to an increase in protein stability of maximally 1.1 kcal mol(-1). The compaction observed is accompanied by increased secondary structure, as far-UV CD spectroscopy shows. Due to the addition of crowding agent, the midpoint of thermal unfolding of native apoflavodoxin rises by 2.9 degrees C. Although the stabilization observed is rather limited, concomitant compaction of unfolded apoflavodoxin restricts the conformational space sampled by the unfolded state, and this could affect kinetic folding of apoflavodoxin. Most importantly, crowding causes severe aggregation of the off-pathway folding intermediate during apoflavodoxin folding in vitro. However, apoflavodoxin can be over expressed in the cytoplasm of Escherichia coli, where it efficiently folds to its functional native form at high yield without noticeable problems. Apparently, in the cell, apoflavodoxin requires the help of chaperones like Trigger Factor and the DnaK system for efficient folding.
Tryptophan-Tryptophan energy migration as a tool to follow apoflavodoxin folding
Visser, N.V. ; Westphal, A.H. ; Hoek, A. van; Mierlo, C.P.M. van; Visser, A.J.W.G. ; Amerongen, H. van - \ 2008
Biophysical Journal 95 (2008). - ISSN 0006-3495 - p. 2462 - 2469.
azotobacter-vinelandii apoflavodoxin - refractive-index - fluorescence depolarization - lipoamide dehydrogenase - glutathione-reductase - flavin fluorescence - hydrogen-exchange - backbone dynamics - protein-structure - flavodoxin-ii
Submolecular details of Azotobacter vinelandii apoflavodoxin (apoFD) (un)folding are revealed by time-resolved fluorescence anisotropy using wild-type protein and variants lacking one or two of apoFD's three tryptophans. ApoFD equilibrium (un)folding by guanidine hydrochloride follows a three-state model: native unfolded intermediate. In native protein, W128 is a sink for Förster resonance energy transfer (FRET). Consequently, unidirectional FRET with a 50-ps transfer correlation time occurs from W167 to W128. FRET from W74 to W167 is much slower (6.9 ns). In the intermediate, W128 and W167 have native-like geometry because the 50-ps transfer time is observed. However, non-native structure exists between W74 and W167 because instead of 6.9 ns the transfer correlation time is 2.0 ns. In unfolded apoFD this 2.0-ns transfer correlation time is also detected. This decrease in transfer correlation time is a result of W74 and W167 becoming solvent accessible and randomly oriented toward one another. Apparently W74 and W167 are near-natively separated in the folding intermediate and in unfolded apoFD. Both tryptophans may actually be slightly closer in space than in the native state, even though apoFD's radius increases substantially upon unfolding. In unfolded apoFD the 50-ps transfer time observed for native and intermediate folding states becomes 200 ps as W128 and W167 are marginally further separated than in the native state. Apparently, apoFD's unfolded state is not a featureless statistical coil but contains well-defined substructures. The approach presented is a powerful tool to study protein folding.
Exploring eddy-covariance and large-aperture scintillometer measurements in an Amazonian rain forest
Randow, C. von; Kruijt, B. ; Holtslag, A.A.M. ; Oliveira, M.B.L. de - \ 2008
Agricultural and Forest Meteorology 148 (2008)4. - ISSN 0168-1923 - p. 680 - 690.
surface-layer fluxes - sensible heat fluxes - refractive-index - optical scintillation - carbon-dioxide - water-vapor - momentum - area - fluctuations - variability
A large-aperture scintillometer (LAS) is used to estimate the surface sensible heat fluxes in an Amazonian rain forest site, and these fluxes are compared with an eddy-covariance system (EC) to analyze conditions of low-frequency modulation in the surface layer. The results show that the flux estimates from the EC are often lower than from the LAS. The differences between EC and LAS tend to increase with decreasing correlation between vertical wind and temperature (rwT). Using different averaging times on EC calculations, we observe that the largest differences between the LAS and the EC fluxes are found for 10-min averages, less so for 30-min averages, while 1-h averages give the smallest differences. The results are attributed to the spatial averaging effect of the LAS. Generally, the results suggest that rwT can be used as an indicator of the importance of low-frequency motions in the surface layer. Evaluating the energy balance for different ranges of rwT, we found that its closure improves when data with increasingly higher rwT are used. In addition, a methodology has been developed to correct the scintillometer signals for the effect of tower vibrations.
Evaluation of two land surface schemes used in terrains of increasing aridity in West Africa
Schüttemeyer, D. ; Moene, A.F. ; Holtslag, A.A.M. ; Bruin, H.A.R. de - \ 2008
Journal of Hydrometeorology 9 (2008)2. - ISSN 1525-755X - p. 173 - 193.
drying semiarid terrain - mesoscale eta-model - water-vapor - canopy conductance - structure parameter - refractive-index - carbon-dioxide - hapex-mobilhy - pine forest - heat-flux
In this study different parameterizations for land surface models currently employed in meteorological models at ECMWF [Tiled ECMWF Surface Scheme for Exchange Processes over Land (TESSEL)] and NCEP (Noah) are evaluated for a semiarid region in Ghana, West Africa. Both schemes utilize the Jarvis¿Stewart approach to calculate canopy conductance as the critical variable for partitioning the available energy into sensible and latent heat flux. Additionally, an approach within Noah is tested to calculate canopy conductance based on plant physiology (A-gs method), where the photosynthetic assimilation is coupled to the leaf stomatal conductance. All parameterizations were run offline for a seasonal cycle in 2002/03 using observations as forcings at two test sites. The two locations are in the humid tropical southern region and in the drier northern region. For the purpose of forcing and evaluation, a new set of data has been utilized to include surface fluxes obtained by scintillometry. The measurements include the rapid wet-to-dry transition after the wet season at both sites. As a general trend, it has been found that during the wet period of a season net radiation is described well by all parameterizations. During the drying process the errors in modeled net radiation increased at both sites. The models perform poorly in simulating soil heat fluxes with larger errors for TESSEL for both sites. The evolution in time for sensible heat flux and latent heat flux was tackled in different ways by the utilized parameterizations and sites with enhanced model performance for the more southern site. Soil moisture in the upper soil layers is modeled with small errors for the different parameterizations. Key adjustments for reducing net radiation during the dry period of a season are discussed. In particular, the ratio of roughness length of momentum and heat was found to be an important parameter, but will require seasonal adjustments
Uncertainty analysis for satellite derived sensible heat fluxes and scintillometer measurements over Savannah environment and comparison to mesoscale meteorological simulation results
Marx, A. ; Kunstmann, H. ; Schüttemeyer, D. ; Moene, A.F. - \ 2008
Agricultural and Forest Meteorology 148 (2008)4. - ISSN 0168-1923 - p. 656 - 667.
land-surface-temperature - large-aperture scintillometer - split-window algorithm - optical scintillation - structure parameter - refractive-index - water-vapor - model - validation - momentum
Three methods for estimating instantaneous sensible heat flux (H) over Savannah environment in West Africa were compared: first, satellite derived estimations using the Surface Energy Balance Algorithm for Land (SEBAL) method [Bastiaanssen, W.G.M., Menenti, M., Feddes, R.A., Holtslag, A.A.M., 1998a. A remote sensing energy balance algorithm for land, SEBAL: 1. Formulation. J. Hydrol. 212¿213, 198¿212]; secondly, measurements at two test sites in Ghana with a large-aperture scintillometer (LAS); third, high resolution mesoscale meteorological simulations using the MM5 (5th-Generation Penn State/NCAR) mesoscale modelling system. Satellite-derived sensible heat flux was based on seven NOAA-16 AVHRR images that were processed for a 2-week period in December 2001 (dry season) and were compared to LAS-data and MM5 simulation results. A methodology based on Gaussian Error Propagation is presented to derive uncertainties in satellite derived sensible heat flux due to (a) input data, (b) coefficients to determine leaf area index (LAI) and (c) methodological differences in estimating surface temperature T0. Total computed relative uncertainty in H was 15% for the Tamale test site and 20% for the Ejura site. Uncertainties in instantaneous evapotranspiration ¿E, however, are much smaller than uncertainties of H. This results due to the same bias in H and Rn ¿ G. For LAS-data, an uncertainty analysis due to input data was performed which showed relative uncertainty of 8% for the Tamale site and 7% for Ejura. Satellite derived net radiation (Rn) was underestimated in comparison to ground measurements which finally caused an underestimation of H. Satellite estimates of H using spatially interpolated ground based measurements of net radiation showed good agreement to LAS data. MM5-computed latent heat flux showed very low values for the entire region. This caused a serious relative MM5-overestimation of sensible heat flux in comparison to LAS and satellite derived estimates. It could be shown that Gaussian Error Propagation can serve as an essential tool to asses the reliability of satellite derived sensible heat fluxes. The resulting uncertainties give information on sensitivities in estimating H and therefore provide a tool for validation purposes.
The use of the scintillation technique for monitoring seasonal water consumption of olive orchards in a semi-arid region
Ezzahar, J. ; Chehbouni, A. ; Hoedjes, J.C.B. ; Er-Raki, S. ; Boulet, G. ; Bonnefonds, J.M. ; Bruin, H.A.R. de - \ 2007
Agricultural Water Management 89 (2007)3. - ISSN 0378-3774 - p. 173 - 184.
large-aperture scintillometer - flevoland field experiment - long-wave-radiation - refractive-index - heterogeneous surface - flux measurements - sonic anemometer - energy fluxes - heat - area
To monitor seasonal water consumption of agricultural fields at large scale, spatially averaged surface fluxes of sensible heat (H) and latent heat (LvE) are required. The scintillation method is shown to be a promising device for obtaining the area-averaged sensible heat fluxes, on a scale of up to 10 km. These fluxes, when combined with a simple available energy model, can be used to derive area-averaged latent heat fluxes. For this purpose, a Large Aperture Scintillometer (LAS) was operated continuously for more than one year over a tall and sparse irrigated oliveyard located in south-central Marrakesh (Morocco). Due to the flood irrigation method used in the site, which induces irregular pattern of soil moisture both in space and time, the comparison between scintillometer-based estimates of daily sensible heat flux (HLAS) and those measured by the classical eddy covariance (EC) method (HEC) showed a large scatter during the irrigation events, while a good correspondence was found during homogenous conditions (dry conditions and days following the rain events). We found, that combining a simple available energy model and the LAS measurements, the latent heat can be reliably predicted at large scale in spite of the large scatter (R2 = 0.72 and RMSE = 18.25 W m¿2) that is obtained when comparing the LAS against the EC. This scatter is explained by different factors: the difference in terms of the source areas of the LAS and EC, the closure failure of the energy balance of the EC, and the error in available energy estimates. Additionally, the irrigation efficiency was investigated by comparing measured seasonal evapotranspiration values to those recommended by the FAO. It was found that the visual observation of the physical conditions of the plant is not sufficient to efficiently manage the irrigation, a large quantity of water is lost (¿37% of total irrigation). Consequently, the LAS can be considered as a potentially useful tool to monitor the water consumption in complex conditions
Hydrometeorological application of a microwave link: 1. Evaporation
Leijnse, H. ; Uijlenhoet, R. ; Stricker, J.N.M. - \ 2007
Water Resources Research 43 (2007). - ISSN 0043-1397 - 9 p.
flevoland field experiment - sensible heat fluxes - heterogeneous surface - refractive-index - large-aperture - scintillometers - fluctuations - absorption - momentum - basin
A method to estimate areal evaporation using a microwave link (radio wave scintillometer) in combination with an energy budget constraint is proposed. This radio wave scintillometry-energy budget method (RWS-EBM) is evaluated for its applicability in different meteorological conditions and for its sensitivity to various variables (the structure parameter of the refractive index of air C n 2, the total available energy R n - G, the wind velocity u, the effective average vegetation height h 0, and the correlation coefficient between the temperature and humidity fluctuations r TQ ). The method is shown to be best suited for use in wet to moderately dry conditions, where the latent heat flux is at least a third of the total available energy (i.e., Bowen ratio =2). It is important to accurately measure the total available energy and the wind velocity as the RWS-EBM is most sensitive to these variables. The Flevoland field experiment has provided the data, obtained with a 27-GHz radio wave scintillometer (over 2.2 km), a large-aperture scintillometer (also 2.2 km), and four eddy covariance systems, which are used to test the RWS-EBM. Comparing 92 daytime measurements (30-min intervals) of the evaporation estimated using the RWS-EBM to that determined in alternative manners (eddy covariance and two-wavelength scintillometry) leads to the conclusion that the method provides consistent estimates (coefficient of determination r 2 = 0.85 in both cases) under relatively wet conditions.