Search PubMed⌕ Search

Biomedical subjects

M J Todd

Publications and source records attributed to M J Todd.

At least 55 records · Page 3Linked to original sources
PubMed1995

The effect of roll-tilt on ocular skew deviation.

Static roll-tilt of normal healthy subjects causes the ocular tilt reaction (OTR) one component of which is disconjugate vertical eye position (skew deviation). In this study the magnitude of skew was measured subjectively by the use of a computerized Hess test at three static roll-tilt angles (head erect, left ear down and right ear down) and two viewing distances (20 cm and 60 cm). The results showed that during static roll-tilt there was a small skew deviation, the magnitude of which was increased at close viewing distances.

Afferent Pathways↗
PubMed1994

Dynamics of the chaperonin ATPase cycle: implications for facilitated protein folding.

The Escherichia coli chaperonins GroEL and GroES facilitate protein folding in an adenosine triphosphate (ATP)-dependent manner. After a single cycle of ATP hydrolysis by the adenosine triphosphatase (ATPase) activity of GroEL, the bi-toroidal GroEL formed a stable asymmetric ternary complex with GroES and nucleotide (bulletlike structures). With each subsequent turnover, ATP was hydrolyzed by one ring of GroEL in a quantized manner, completely releasing the adenosine diphosphate and GroES that were tightly bound to the other ring as a result of the previous turnover. The catalytic cycle involved formation of a symmetric complex (football-like structures) as an intermediate that accumulated before the rate-determining hydrolytic step. After one to two cycles, most of the substrate protein dissociated still in a nonnative state, which is consistent with intermolecular transfer of the substrate protein between toroids of high and low affinity. A unifying model for chaperonin-facilitated protein folding based on successive rounds of binding and release, and partitioning between committed and kinetically trapped intermediates, is proposed.

Adenosine Triphosphatases↗
PubMed1994

On the role of groES in the chaperonin-assisted folding reaction. Three case studies.

The mechanism by which correctly folded proteins are recovered from stable complexes with groEL is not well understood. Certain target proteins require ATP and groES, while others seemingly dispense with the cochaperonin. Here, we examine the chaperonin-assisted folding of ribulose-1,5-bisphosphate carboxylase, malate dehydrogenase, and citrate synthase, three proteins that are believed to require both chaperonin components for successful reactivation. Surprisingly, in all cases, the need for groES depended on the folding environment. Under "non-permissive" conditions, where unassisted spontaneous folding could not occur, reactivation to the native state required the complete chaperonin system (e.g. groEL, groES, and MgATP). However, under "permissive" conditions where spontaneous folding could occur groES was no longer mandatory. Instead, upon the addition of ATP alone, all three target proteins could be released from groEL, in a form that was capable of reaching the native state. In the permissive setting, groES merely accelerated the rate of the ATP-dependent release process. The results suggest that the incompletely folded protein species that are released from groEL, in the absence of groES, are not necessarily committed to the native state. Similar to the unassisted folding reaction, they still partition between productive and unproductive folding pathways in an environment-dependent manner. It follows that the mechanistic contribution of the co-chaperonin, groES, and its physiological significance in cellular protein folding, could be entirely missed in a permissive in vitro environment.

Animals↗
PubMed1994

Unilateral vestibular deafferentation causes permanent impairment of the human vertical vestibulo-ocular reflex in the pitch plane.

Rapid, passive, unpredictable, low-amplitude (10-20 degrees), high-acceleration (3000-4000 degrees/s2) head rotations were used to study the vertical vestibulo-ocular reflex in the pitch plane (pitch-vVOR) after unilateral vestibular deafferentation. The results from 23 human subjects who had undergone therapeutic unilateral vestibular deafferentation were compared with those from 19 normals. All subjects were tested while seated in the upright position. Group means and two-tailed 95% confidence intervals are reported for the pitch-vVOR gains in normal and unilateral vestibular deafferented subjects. In normal subjects, at a head velocity of 125 degrees/s the pitch-vVOR gains were: upward 0.89 +/- 0.06, downward 0.91 +/- 0.04. At a head velocity of 200 degrees/s, the pitch-vVOR gains were: upward 0.92 +/- 0.06, downward 0.96 +/- 0.04. There was no significant up-down asymmetry. In the 15 unilateral vestibular deafferented subjects who were studied more than 1 year after unilateral vestibular deafferentation, the pitch-vVOR was significantly impaired. At a head velocity of 125 degrees/s, the pitch-vVOR gains were: upward 0.67 +/- 0.11, downward 0.63 +/- 0.07. At a head velocity of 200 degrees/s, the pitch-vVOR gains were: upward 0.67 +/- 0.07, downward 0.58 +/- 0.06. There was no significant up-down asymmetry. The pitch-vVOR gain in unilateral vestibular deafferented subjects was significantly lower (P < 0.05) than the pitch-vVOR gain in normal subjects at the same head velocities. These results show that total, permanent unilateral loss of vestibular function produces a permanent symmetrical 30% (approximately) decrease in pitch-vVOR gain. This pitch-vVOR deficit is still present more than 1 year after deafferentation despite retinal slip velocities greater than 30 degrees/s in response to head accelerations in the physiological range, indicating that compensation of pitch-vVOR function following unilateral vestibular deafferentation remains incomplete.

Adult↗
PubMed1994

Jerk-waveform see-saw nystagmus due to unilateral meso-diencephalic lesion.

See-saw nystagmus is an uncommon but highly characteristic eye movement disorder comprising intorsion and elevation of one eye, with synchronous extorsion and depression of the other. It generally has a pendular waveform and is due to a midline, extrinsic, suprasellar mass lesion compressing or invading the brainstem bilaterally at the meso-diencephalic junction. This report deals with the clinical and MRI findings in three patients (and binocular three-dimensional quantitative oculographic findings in one patient) with a jerk waveform see-saw nystagmus due in each case to a unilateral meso-diencephalic lesion. In each patient the torsional component of the nystagmus fast phases rotated the upper poles of the eyes toward the side of the lesion. Jerk see-saw nystagmus can be clinically indistinguishable from pendular see-saw nystagmus and from the torsional-vertical nystagmus which occurs with medullary lesions. We propose that jerk see-saw nystagmus is due to unilateral inactivation of the torsional eye-velocity integrator, thought to be in the interstitial nucleus of Cajal, with sparing of the torsional fast-phase generator, thought to be in the adjacent rostral interstitial nucleus of the medial longitudinal fasciculus.

Adult↗
PubMed1993

Identification and functional analysis of chaperonin 10, the groES homolog from yeast mitochondria.

Chaperonin 60 (cpn60) and chaperonin 10 (cpn10) constitute the chaperonin system in prokaryotes, mitochondria, and chloroplasts. In Escherichia coli, these two chaperonins are also termed groEL and groES. We have used a functional assay to identify the groES homolog cpn10 in yeast mitochondria. When dimeric ribulose-1,5-bisphosphate carboxylase (Rubisco) is denatured and allowed to bind to yeast cpn60, subsequent refolding of Rubisco is strictly dependent upon yeast cpn10. The heterologous combination of cpn60 from E. coli plus yeast cpn10 is also functional. In contrast, yeast cpn60 plus E. coli cpn10 do not support refolding of Rubisco. In the presence of MgATP, yeast cpn60 and yeast cpn10 form a stable complex that can be isolated by gel filtration and that facilitates refolding of denatured Rubisco. Although the potassium-dependent ATPase activity of E. coli cpn60 can be inhibited by cpn10 from either E. coli or yeast, neither of these cpn10s inhibits the ATPase activity of yeast cpn60. Amino acid sequencing of yeast cpn10 reveals substantial similarity to the corresponding cpn10 proteins from rat mitochondria and prokaryotes.

Adenosine Triphosphatases↗
PubMed1993

Hydrolysis of adenosine 5'-triphosphate by Escherichia coli GroEL: effects of GroES and potassium ion.

The potassium-ion activation constant (Kact) for the ATPase activity of Escherichia coli chaperonin groEL is inversely dependent upon the ATP concentration over at least 3 orders of magnitude. The ATPase activity shows positively cooperative kinetics with respect to ATP and K+. Both the K0.5 for ATP and cooperativity (as measured by the Hill coefficient) decrease as the K+ concentration increases. Equilibrium binding studies under conditions where hydrolysis does not occur indicate that MgATP binds weakly to groEL in the absence of K+. In the absence of groES, the K(+)-dependent hydrolysis of ATP by groEL continues to completion. In the presence of groES, the time course for the hydrolysis of ATP by groEL becomes more complex. Three distinct kinetic phases can be discerned. Initially, both heptameric toroids turn over once at the same rate that they do in the absence of groES. This leads to the formation of an asymmetric binary complex, groEL14-MgADP7-groES7, in which 7 mol of ADP is trapped in a form that does not readily exchange with free ADP. In the second phase, the remaining seven sites (containing readily exchangeable ADP) turn over, or have the potential to turn over, at the same rate as they do in the absence of groES, so that the overall rate of hydrolysis is maximally 50%. These remaining sites of the asymmetric binary complex do not hydrolyze all of the available ATP. Instead, the second phase of hydrolysis gives way to a third, completely inhibited state, the onset of which is dependent upon the relative affinities of the remaining sites for MgATP and MgADP.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Diphosphate↗
PubMed1993

Chaperonins and protein folding: unity and disunity of mechanisms.

Chaperonin-facilitated folding of proteins involves two partial reactions. The first partial reaction, the formation of stable binary complexes between chaperonin-60 and non-native states of the target protein, is common to the chaperonin-facilitated folding of all target proteins investigated to date. The structural basis for this interaction is not presently understood. The second partial reaction, the dissociation of the target protein in a form committed to the native state, appears to proceed by a variety of mechanisms, dependent upon the nature of the target protein in question. Those target proteins (e.g. rubisco, rhodanese, citrate synthase) which require the presence of chaperonin-10, also appear to require the hydrolysis of ATP to bring about the dissociation of the target protein from chaperonin-60. With one exception (pre-beta-lactamase) those target proteins which do not require the presence of chaperonin-10 to be released from chaperonin-60, also do not require the hydrolysis of ATP, since non-hydrolysable analogues of ATP support the release of the target protein in a state committed to the native state. The question of whether or not chaperonin-facilitated folding constitutes a catalysed event is addressed.

Animals↗
PubMed1991

Identification of the essential cysteine residue in Klebsiella aerogenes urease.

During reaction with [14C]iodoacetamide at pH 6.3, radioactivity was incorporated primarily into a single Klebsiella aerogenes urease peptide concomitant with activity loss. This peptide was protected from modification at pH 6.3 by inclusion of phosphate, a competitive inhibitor of urease, which also protected the enzyme from inactivation. At pH 8.5, several peptides were alkylated; however, modification of one peptide, identical to that modified at pH 6.3, paralleled activity loss. The N-terminal amino acid sequence and composition of the peptide containing the essential thiol was determined. Previous enzyme inactivation studies of K. aerogenes urease could not distinguish whether one or two essential thiols were present per active site (Todd, M. J., and Hausinger, R. P. (1991) J. Biol. Chem. 266, 10260-10267); we conclude that there is a single essential thiol present and identify this residue as Cys319 in the large subunit of the heteropolymeric enzyme.

Amino Acid Sequence↗
PubMed1991

Reactivity of the essential thiol of Klebsiella aerogenes urease. Effect of pH and ligands on thiol modification.

The kinetics of Klebsiella aerogenes urease inactivation by disulfide and alkylating agents was examined and found to follow pseudo-first-order kinetics. Reactivity of the essential thiol is affected by the presence of substrate and competitive inhibitors, consistent with a cysteine located proximal to the active site. In contrast to the results observed with other reagents, the rate of activity loss in the presence of 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) saturated at high reagent concentrations, indicating that DTNB must first bind to urease before inactivation can occur. The pH dependence for the rate of urease inactivation by both disulfide and alkylating agents was consistent with an interaction between the thiol and a second ionizing group. The resulting macroscopic pKa values for the 2 residues are less than 5 and 12. Spectrophotometric studies at pH 7.75 demonstrated that 2,2'-dithiodipyridine (DTDP) modified 8.5 +/- 0.2 mol of thiol/mol of enzyme or 4.2 mol of thiol/mol of catalytic unit. With the slow tight binding competitive inhibitor phenyl-phosphorodiamidate (PPD) bound to urease, 1.1 +/- 0.1 mol of thiol/mol of catalytic unit were protected from modification. PPD-bound DTDP-modified urease could be reactivated by dialysis, consistent with the presence of one thiol per active site. Analogous studies at pH 6.1, using the competitive inhibitor phosphate, confirmed the presence of one protected thiol per catalytic unit. Under denaturing conditions, 25.5 +/- 0.3 mol of thiol/mol of enzyme (Mr = 211, 800) were modified by DTDP.

2,2'-Dipyridyl↗
PubMed1990

The human horizontal vestibulo-ocular reflex in response to high-acceleration stimulation before and after unilateral vestibular neurectomy.

The normal horizontal vestibulo-ocular reflex (HVOR) is largely generated by simultaneous stimulation of the two horizontal semicircular canals (HSCCs). To determine the dynamics of the HVOR when it is generated by only one HSCC, compensatory eye movements in response to a novel vestibular stimulus were measured using magnetic search coils. The vestibular stimulus consisted of low-amplitude, high-acceleration, passive, unpredictable, horizontal rotations of the head with respect to the trunk. While these so called head "impuses" had amplitudes of only 15-20 degrees with peak velocities up to 250 deg/s, they had peak accelerations up to 3000 deg/s/s. Fourteen humans were studied in this way before and after therapeutic unilateral vestibular neurectomy; 10 were studied 1 week or 1 year afterwards; 4 were studied 1 week and 1 year afterwards. The results from these 14 patients were compared with the results from 30 normal control subjects and with the results from one subject with absent vestibular function following bilateral vestibular neurectomy. Compensatory eye rotation in normal subjects closely mirrored head rotation. In contrast there was no compensatory eye rotation in the first 170 ms after the onset of head rotation in the subject without vestibular function. Before unilateral vestibular neurectomy all the patients' eye movement responses were within the normal control range. One week after unilateral vestibular neurectomy however there was a asymmetrical bilateral HVOR deficit. The asymmetry was much more profound than has been shown in any previous studies. The HVOR generated in response to head impulses directed away from the intact side largely by ampullofugal disfacilitation from the single intact HSCC (ignoring for the moment the small contribution to the HVOR from stimulation of the vertical SCCs), was severely deficient with an average gain (eye velocity/head velocity) of 0.25 at 122.5 deg/sec head velocity (normal gain = 0.94 +/- 0.08). In contrast the HVOR generated in response to head impulses directed toward the intact side, largely by ampullopetal excitation from the single intact HSCC, was only mildly (but nonetheless significantly) deficient, with an average gain of 0.80 at 122.5 deg/sec head velocity. At these accelerations there was no significant improvement in the average HVOR velocity gain in either direction over the following year. These results indicate that ampullopetal excitation from one HSCC can, even in the absence of ampullofugal disfacilitation from the opposite HSCC, generate a near normal HVOR in response to high-acceleration stimulation.(ABSTRACT TRUNCATED AT 400 WORDS)

Acceleration↗
PubMed1989

Competitive inhibitors of Klebsiella aerogenes urease. Mechanisms of interaction with the nickel active site.

We examined several compounds for their mechanisms of inhibition with the nickel-containing active site of homogeneous Klebsiella aerogenes urease. Thiolate anions competitively inhibit urease and directly interact with the metallocenter, as shown by the pH dependence of inhibition and by UV-visible absorbance spectroscopic studies. Cysteamine, which possesses a cationic beta-amino group, exhibited a high affinity for urease (Ki = 5 microM), whereas thiolates containing anionic carboxyl groups were uniformly poor inhibitors. Phosphate monoanion competitively inhibits a protonated form of urease with a pKa of less than 5. Both the thiolate and phosphate inhibition results are consistent with charge repulsion by an anionic group in the urease active site. Acetohydroxamic acid (AHA) was shown to be a slow-binding competitive inhibitor of urease. This compound forms an initial E.AHA complex which then undergoes a slow transformation to yield an E.AHA* complex; the overall dissociation constant of AHA is 2.6 microM. Phenylphosphorodiamidate, also shown to be a slow-binding competitive inhibitor, possesses an overall dissociation constant of 94 pM. The tight binding of phenylphosphorodiamidate was exploited to demonstrate the presence of two active sites per enzyme molecule. Urease contains 4 mol of nickel/mol enzyme, hence there are two nickel ions/catalytic unit. Each of the two slow-binding inhibitors are proposed to form complexes in which the inhibitor bridges the two active site nickel ions. The inhibition results obtained for K. aerogenes urease are compared with inhibition studies of other ureases and are interpreted in terms of a model for catalysis proposed for the jack bean enzyme (Dixon, N.E., Riddles, P.W., Gazzola, C., Blakely, R.L., and Zerner, B. (1980) Can. J. Biochem. 58, 1335-1344).

Binding Sites↗