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A T Smith

Publications and source records attributed to A T Smith.

At least 73 records · Page 4Linked to original sources
PubMed1995

Crystallization and preliminary X-ray studies of recombinant horseradish peroxidase.

A non-glycosylated form of horseradish peroxidase c extracted from Escherichia coli inclusion bodies and refolded in the presence of haem and Ca(2+) ions has been used to grow protein crystals suitable for X-ray diffraction analysis. The crystals are prisms in the trigonal space group P3(1)12 or P3(2)12 with a = b = 158.9 and c = 114.3 A, and diffract to 1.9 A. There are four molecules, each of 34 kDa, in the asymmetric unit. The molecules of the asymmetric unit are related by approximate translational symmetry, resulting in pseudo-centerings. Data to approximately 15 A can thus be described by a lattice of a' = b' = 91.7 A and c' = 57.1 A, alpha = beta = 90 degrees and gamma = 120 degrees, including four molecules.

Journal Article↗
PubMed1995

Sharpening of drifting, blurred images.

The perceived blur of moving images is less than expected given the sluggish temporal response of the visual system. This suggests that a motion deblurring mechanism may exist to preserve the positional acuity and sharpness of moving images. Furthermore, when sequences of blurred stills are presented, observers report that the moving image is in sharp focus raising the possibility that there is a mechanism which may sharpen the appearance of moving, blurred images. We have measured the effects of velocity and contrast on the perceived blur of drifting, blurred images (sine gratings and blurred edges). Subjects matched the perceived blur of drifting, blurred images to that of static, blurred images in a dimly lit room. It was found that perceived blur was inversely related to drift speed and contrast. The results confirm that moving, blurred images may appear sharper than when they are static. This finding is not consistent with some models of motion deblurring since these account only for the preservation of sharp contours that are present in the image and not for the sharp appearance of images that are in fact blurred.

Contrast Sensitivity↗
PubMed1995

The perceived speed of second-order motion and its dependence on stimulus contrast.

Speed matches were obtained, using a spatial two-alternative forced-choice task, between a second-order motion stimulus and a first-order motion stimulus. The second-order motion stimulus was composed of contrast-modulated noise [produced by multiplying two-dimensional (2-d), static noise by a drifting, one-dimensional (1-d) sinusoid]. The first-order motion stimulus was composed of luminance-modulated noise (produced by summing, rather than multiplying, 2-d noise and a drifting sine grafting). In Expt 1, the relationship between the perceived speed of first- and second-order motion was examined. The motion stimuli had the same spatial frequency (1 or 3 c/deg) and were equated for visibility by presenting them at the same multiple of direction-identification threshold. Over a range of physical speeds, the perceived speeds of the first-order and second-order motion stimuli were identical when their physical speeds were the same. In Expt 2, the effect of varying stimulus "contrast" (contrast modulation depth) on the perceived speed of second-order motion was examined. The contrast of the first-order motion stimulus was fixed and speed matches were obtained for second-order motion stimuli at several contrast modulation depths. The motion stimuli had the same spatial (1 or 4 c/deg) and temporal (5 or 20 Hz) frequencies. It was found that the perceived speed of second-order motion was approximately linearly related to log modulation depth. In agreement with previous studies we also confirmed that the perceived speed of first-order motion is similarly dependent on stimulus contrast (luminance modulation depth).(ABSTRACT TRUNCATED AT 250 WORDS)

Contrast Sensitivity↗
PubMed1994

pH-dependent properties of a mutant horseradish peroxidase isoenzyme C in which Arg38 has been replaced with lysine.

Arg38 in the active site of horseradish peroxidase isoenzyme C (HRP-C) has been replaced with lysine by site-directed mutagenesis. As a preclude to a detailed kinetic analysis of this variant, the present study characterizes a pH-dependent cycle of reactions for recombinant horseradish peroxidase isoenzyme C with Arg38 replaced by lysine ([R38K]HRP-C*), which involves time-dependent changes in both specific activity and the electronic absorption spectrum of the enzyme. This pH-dependent cycle resembles that previously suggested for a cytochrome-c peroxidase variant in which Asp235 was replaced with asparagine. When the pH of a solution of resting [R38K]HRP-C* at pH 6.6 (form AH) is raised to pH 8.6, a rapid alkaline transition occurs. This results in spectral changes characteristic of a shift from a predominantly pentacoordinate to a completely hexacoordinate high-spin haem iron (form A-) with a pKa of 7.5. When the pH of a solution of form A- is raised from 8.7 to 12.0, no further spectral changes are observed. The reaction is reversible, but when the high-pH form of the enzyme (A-) is allowed to stand at pH 8.6, it slowly becomes converted into a third enzyme form (form I-) at a rate which is independent of pH (k = 0.56 h-1). When the pH of a sample of form I- is lowered from 8.6 to 6.6, the original low-pH form (AH) of the enzyme is recovered. Recovery of form AH from form I- does not occur via form A-, but via at least one further intermediate, form X. Following a downward pH jump, the rate constant for the formation of form X from form I- shows a small dependence on pH, changing from 48 s-1 at pH 6.8 to 39 s-1 at pH 7.4. The rate of formation of form AH from form X is also pH dependent and biphasic in nature, with measured rate constants ranging from 11.9-2.1 h-1. The possible structures of the different forms of [R38K]HRP-C* are discussed in the light of similar data in the literature for variants of cytochrome-c peroxidase. The properties may be indicative of a greater degree of conformational flexibility within the active site of this mutant caused by the smaller bulk of the lysine side-chain and the probable disruption of a part of the haem-linked hydrogen-bonding network in the distal haem pocket. The wild-type enzyme undergoes no such pH induced changes.

Arginine↗
PubMed1994

Characterization of recombinant horseradish peroxidase C and three site-directed mutants, F41V, F41W, and R38K, by resonance Raman spectroscopy.

Resonance Raman spectra are reported for recombinant horseradish peroxidase C (HRP-C*) and three protein variants prepared by in vitro refolding after Escherichia coli expression. The spectra of their FeII and FeIII forms and of their complexes with benzohydroxamic acid (BHA) were recorded at neutral pH. The residues mutated were on the distal [Phe41-->Trp or Val (F41W, F41V) and Arg38-->Lys (R38K)] side of the heme. The spectra give information on the spin and ligation states via the frequencies of the core size marker bands. No detectable modification in the enzyme structure or in the heme group has been observed in the wild-type recombinant HRP-C*. The FeIII forms of both the recombinant and the plant proteins show the coexistence of a 5-(5-cHS) and a 6-coordinate high-spin (6-cHS) heme, characterized by the anomalous frequencies of certain bands, namely, v3 and v10, which we attribute to a different degree of distortion of the heme planarity with respect to other heme proteins and model compounds, resulting from external forces such as steric contacts within the protein. This effect is partially relieved upon complexation with BHA or as a result of mutation. F41W and F41V are characterized by an increase in a 6-cHS form at the expense of the 5-cHS species, and the R38K by an increase in both the 6-c high-(HS) and low-spin (LS) hemes. The 6-cHS and -LS species are characterized by normal core size marker band frequencies. The FeII-His RR band is at 243 cm-1 in HRP-C*, the high frequency value being due to hydrogen-bonding interactions between the proximal His170 N delta and the carboxylate acceptor group on Asp247. Mutation at position 38 causes a downshift of 3 cm-1 in the v(Fe-Im) stretching mode, suggesting a weakening of the Fe-Im bond strength. By comparing the results obtained with HRP-C* mutants with those previously reported for the corresponding cytochrome c peroxidase (CCP) mutants, it appears that the distal heme pocket architecture is significantly different in the two peroxidases, although the hydrogen-bonding network coupling the distal and the proximal sides of the heme appears to be conserved. Mutations on the distal side dramatically affect the capability of the protein to bind BHA. F41W and R38K mutants do not bind the substrate, whereas the F41V variant shows a 2-fold increase in affinity.(ABSTRACT TRUNCATED AT 400 WORDS)

Base Sequence↗
PubMed1994

Temporal beats in the human visual system.

The human visual system is generally assumed to comprise multiple independent channels tuned to limited ranges of spatial and temporal frequency. However, it is known that spatial beats effectively mask a sinusoidal grating whose spatial frequency coincides with the beat frequency of the mask, even though no Fourier energy exists in the mask at that frequency. We investigated whether analogous masking effects occur in the temporal domain and whether adaptation to either spatial or temporal beats results in threshold elevation. Our results show that temporal beat patterns do not mask a grating temporally modulated at the beat frequency. Adaptation to neither spatial nor temporal beats results in measurable threshold elevation at the beat frequency. Since a luminance non-linearity would lead to temporal as well as spatial distortion products, we conclude that sensitivity to spatial beats does not rely on the presence of distortion products but instead reflects subsequent processing that occurs in the spatial domain only. The most likely site of such processing is cortical.

Adaptation, Ocular↗
PubMed1994

Evidence for separate motion-detecting mechanisms for first- and second-order motion in human vision.

Current theories of second-order motion perception postulate that such motion is detected by either a high-level mechanism which computes the temporal correspondences between "features" extracted from the image, or low-level motion mechanisms which operate on a nonlinear, neural transformation of the luminance profile of the image. Theories which favour the latter strategy either suggest that first- and second-order motion are detected by a common mechanism or else that distinct mechanisms exist for the two types of motion, both operating on similar principles. The aim of this study was to differentiate between these possibilities. Observers were required to judge the direction of multiframe motion sequences in which the frames alternated between sinusoidal variations in luminance (first order) and similar variations in contrast (second order). On each frame the modulation signal was displaced by some fraction of its spatial period. The motion sequences were designed such that integration of both types of frame (first and second order) would lead to unambiguous motion in a particular direction whilst separate analysis of first- or second-order frames alone would yield ambiguous motion. The results show clearly that observers were unable to integrate the first- and second-order frames of such motion sequences. However, when observers were presented with motion sequences in which the frames alternated between two, different types of second-order image (variations in the contrast or size of the elements constituting a random noise field) perceived direction was always consistent with integration of both image types. This is taken as support for models that suggest that first- and second-order motion are processed by distinct mechanisms in the visual system and that each mechanism is only sensitive to one type of motion. It is suggested that several varieties of second-order motion stimuli may be regarded as equivalent to contrast-modulated images when considered in terms of the effects of local spatiotemporal filtering operations carried out by the human visual system. In this respect, our results are consistent with the "texture grabber" concept of Werkhoven, Sperling and Chubb [(1993) Vision Research, 33, 463-485].

Contrast Sensitivity↗
PubMed1994

Perceived contrast as a function of adaptation duration.

We measured how the perceived contrast of a sinusoidal grating fades as a function of time. Measurements were made for a range of temporal and spatial frequencies and eccentricities. Patterns of high temporal and low spatial frequency exhibited a greater and more rapid loss of apparent contrast (fade) than those of medium frequencies. The rate and amount of fading for a subgroup of moderate frequencies increased when presented peripherally rather than foveally. Further measurements revealed that gratings of disparate spatial frequencies, but with the same threshold sensitivity, exhibit very different fading characteristics but equal threshold elevation. We conclude that the differential loss of apparent contrast is not an artefact of differing proximities to threshold, nor can it be accounted for by differences in the adaptability of underlying spatio-temporal mechanisms at threshold. The differences in fading may thus reflect either a difference in the adaptability of underlying channels above threshold or a differential contribution of such channels to perceived contrast.

Adaptation, Ocular↗
PubMed1994

Is global motion really based on spatial integration of local motion signals?

Previous studies have shown that a random-dot kinematogram (RDK) comprising dots, each of which takes a random walk in direction or speed over time, can appear to flow in a single direction. This has been interpreted as evidence for the existence of a co-operative network linking neurons sensitive to different directions/speeds and different spatial locations. We have investigated the possibility that global motion perception in such patterns might simply reflect motion energy detection at a coarse spatial scale (such that many dots fall in the receptive field of one energy detector) without the need to encode local dot motions on a fine spatial scale and then integrate their motions over space. We created random-walk RDKs and then spatially high-pass filtered them to remove low spatial frequencies. Perception of global motion was unimpaired for both direction and speed random walks, showing that the phenomenon is not reliant on low spatial frequencies and must, therefore, involve integration of local motion signals across space, as originally postulated.

Discrimination, Psychological↗
PubMed1994

Antagonistic comparison of temporal frequency filter outputs as a basis for speed perception.

The prevailing view of motion detection in human vision is that the retinal image is convolved with each of a set of spatiotemporal filters and that perceived speed emerges from a process of pooling the outputs of these filters. Such a system can operate only if multiple filters exist; ideally the filters should also be fairly narrowly tuned in both spatial and temporal frequency. These constraints are met in the case of spatial frequency. But several studies suggest that multiple, finely tuned temporal filters do not exist; instead just two (perhaps three) broadly-tuned temporal mechanisms can be identified. We report some experiments concerning the effects of adaptation to motion on perceived speed. It is shown that perceived speed is increased by adaptation in some circumstances and decreased in others. We then present a computational model in which a temporal frequency code, on which perceived speed is presumed to be based, is derived by a process of antagonistic comparison of the responses of two psychophysically-plausible, broadly-tuned temporal mechanisms. The model, which includes the effects of adaptation to motion upon the sensitivities of the filters and the subsequent comparison of their sensitivities, is shown to give a good fit to the empirical data.

Adaptation, Ocular↗
PubMed1994

The duration of the motion aftereffect following adaptation to first-order and second-order motion.

The magnitude of the motion aftereffect (MAE) obtained following adaptation to first-order or to second-order motion was measured by estimating its duration. The second-order adaptation stimulus was composed of contrast-modulated noise produced by multiplying two-dimensional (2-D) noise by a drifting 1 cycle deg-1 sine grating. The first-order adaptation stimulus was composed of luminance-modulated noise produced by summing, rather than multiplying, the noise and the sine grating. The test stimuli were directionally ambiguous motion patterns composed of either two oppositely drifting sine gratings added to noise or the contrast-modulated equivalent. The adaptation and test stimuli were equated for visibility by presenting them at the same multiple of direction-identification threshold. All possible combinations of first-order and second-order adaptation and test stimuli were examined in order to compare the magnitudes of the MAEs obtained following same adaptation and cross adaptation. After adaptation the test stimuli always appeared to drift coherently in the direction opposite to that of adaptation and the magnitudes of this MAE were very similar for all conditions examined. Statistical analyses of the results showed that there was no significant difference between the durations of the MAEs obtained in the same-adaptation and cross-adaptation conditions. The cross-adaptation effects suggest that either first-order or second-order motion are detected by a common low-level mechanism, or that separate parallel motion-detecting mechanisms exist, for the two types of motion, that interact at some later stage of processing.

Humans↗
PubMed1994

Direction identification thresholds for second-order motion in central and peripheral vision.

Evidence bearing on the question of whether first-order and second-order motion are detected by use of the same or different principles has been sought. This question was approached by measuring thresholds for correctly identifying the direction of motion of various second-order motion patterns. The patterns used were contrast-modulated noise patterns in which the contrast of a carrier was modulated sinusoidally in one dimension, and the modulating waveform drifted smoothly while the carrier itself remained stationary. The carrier used was in most cases static two-dimensional noise; other carriers gave similar results. Thresholds were measured in terms of amplitude of contrast modulation (modulation depth) for each of a range of envelope drift speeds and spatial frequencies in the fovea and at several viewing eccentricities. Along with direction-identification thresholds, thresholds for either simple detection of the modulation or for correctly identifying the orientation of the modulation were simultaneously measured. Thresholds for direction identification were generally somewhat higher than those for simple detection. However, they were in most cases very similar to thresholds for identification of orientation, as found for conventional luminance gratings. Contrary to some reports, sensitivity to contrast-modulated patterns declines with eccentricity at a similar rate to that found with luminance gratings. The results suggest that first-order and second-order motion are either detected by a common motion-detection mechanism or are detected by different mechanisms that use a common principle of motion detection.

Humans↗
PubMed1994

Correspondence-based and energy-based detection of second-order motion in human vision.

Motion in the retinal image may occur either in the form of spatiotemporal variations in luminance (first-order motion) or as spatiotemporal variations in characteristics derived from luminance, such as contrast (second-order motion). Second-order motion patterns were employed in an attempt to establish the principles used for the detection of image motion in the human visual system. In principle, one can detect motion at a high level of visual analysis by identifying features of the image and tracking their positions (correspondence-based detection) or at a low level by analysis of spatiotemporal luminance variations without reference to features (intensity-based detection). Prevailing models favor the latter approach, which has been adapted to account for the visibility of second-order motion by postulation of a stage of rectification that precedes motion energy detection [J. Opt. Soc. Am. A 5, 1986 (1988)]. In two experiments it is shown that second-order motion is indeed detected normally by use of the strategy of transformation plus energy detection but that detection can also be achieved by use of the feature-correspondence strategy when the intensity strategy fails. In the first experiment, a stimulus is employed in which opposite directions of motion perception are predicted by the two strategies. It is shown that normally the direction associated with motion energy in the rectified image is seen but that the direction associated with feature motion is seen when the energy system is disabled by the use of an interstimulus interval.(ABSTRACT TRUNCATED AT 250 WORDS)

Contrast Sensitivity↗
PubMed1993

Transparent motion from feature- and luminance-based processes.

The apparent motion of a discretely displaced complex waveform with a periodic contrast modulation or "beat" of frequency f and sinusoidal components of frequencies 3f and 4f was examined at various interstimulus intervals (ISIs). At short ISIs perception of motion of both the beat and an aliased component of the waveform results in transparent motion. At longer ISIs motion is perceived only in the direction of the features of the waveform. The transparent motion observed at short ISIs indicates that, under certain conditions, "short-range" motion sensors do not constrain "long-range" feature processing and both may be active simultaneously.

Adaptation, Ocular↗
PubMed1993

The use of orthodontic chain elastic for temporary intermaxillary fixation.

A method of application of firm and resilient temporary intermaxillary fixation using readily available orthodontic elastomeric chain is described. Advantages lie in the time efficiency of its use, relative safety compared to tie-wiring techniques, flexibility of direction of pull, and the ease and rapidity with which it can be removed. Consideration is given to other potential uses of the material in the retention of various appliances used in orthognathic procedures.

Equipment Design↗
PubMed1992

Characterisation of a haem active-site mutant of horseradish peroxidase, Phe41----Val, with altered reactivity towards hydrogen peroxide and reducing substrates.

A horseradish peroxidase variant ([F41V] HRP-C*), in which Val replaces the conserved Phe at position 41 adjacent to the distal His, has been constructed. Its composition and spectroscopic, catalytic and substrate-binding properties were compared with those of the wild-type recombinant (HRP-C*) and plant (HRP-C) enzymes. Presteady-state kinetic measurements of the rate constant for compound I formation (k1) revealed an eightfold decrease in the reactivity of the Phe41----Val variant towards H2O2, in comparison with HRP-C or HRP-C*. Measurement of the remaining rate constants, k2 and k3, for the two single-electron reduction reactions of [F41V] HRP-C with para-aminobenzoic acid as reducing substrate, showed that they were 2.5-fold and 1.3-fold faster, respectively. In contrast, analysis of data from steady-state assays with 2,2'-azinobis(3-ethylbenzthiazoline-6-sulphonate) as reducing substrate, showed decreased reactivity of the mutant enzyme to this compound, indicating a change in substrate specificity. Over the substrate range studied, the data for HRP-C* and for [F41V] HRP-C conformed to a simple modification of the accepted peroxidase mechanism in which a first-order step (ku), assumed to be product dissociation, becomes rate-limiting under our standard assay conditions. Calculations of rate constants from steady-state data yielded values of k1 for both enzyme forms in adequate agreement with those from pre-steady state measurements. They showed, furthermore, that both k3 for 2,2'-azinobis(3-ethylbenzthiazoline-6-sulphonate) and ku were substantially decreased, fivefold and tenfold, respectively, in the mutant. Analogous to the decrease in ku, we observed a twofold increase in the affinity of the mutant variant for the inhibitor benzhydroxamic acid. The coordination-state equilibrium of the haem iron also appeared shifted towards the hexacoordinate high-spin form. These observations indicate that in addition to affecting reactivity to H2O2, mutations in the distal region and close to the haem iron also affect reactivity towards different reducing substrates, inducing perturbations in the neighbourhood of the aromatic-substrate-binding site, known to be 0.8-1.2 nm from the haem iron.

Amino Acid Sequence↗