PubMed Health⌕ Search

Biomedical subjects

J McCracken

Publications and source records attributed to J McCracken.

At least 19 recordsLinked to original sources
PubMed2003

Bovine parotid secretory protein: structure, expression and relatedness to other BPI (bactericidal/permeability-increasing protein)-like proteins.

Members of the family of BPI (bactericidal/permeability-increasing protein)-like proteins are as yet incompletely characterized, particularly in cattle, where full-length sequence information is available for only three of the 13 family members known from other species. Structural bioinformatic analyses incorporating bovine homologues of several members of the BPI-like protein family, including two forms of bovine parotid secretory protein (PSP), showed that this family is also present in cattle. Expression analyses of several members of the BPI-like protein family in cattle, including PSP (Bsp30), von Ebner's minor salivary gland protein (VEMSGP) and lung-specific X protein (LUNX), showed a restricted pattern of expression, consistent with earlier hypotheses that these proteins function in the innate immune response to bacteria. The possible role of bovine PSP in susceptibility to pasture bloat in cattle is discussed.

Animals↗
PubMed2001

Fast deuterium access to the buried magnesium/manganese site in cytochrome c oxidase.

The recently determined crystal structures of bacterial and bovine cytochrome c oxidases show an area of organized water within the protein immediately above the active site where oxygen chemistry occurs. A pathway for exit of protons or water produced during turnover is suggested by possible connections of this aqueous region to the exterior surface. A non-redox-active Mg(2+) site is located in the interior of this region, and our previous studies [Florens, L., Hoganson, C., McCracken, J., Fetter, J., Mills, D., Babcock, G. T., and Ferguson-Miller, S. (1998) in Phototropic Prokaryotes (Peschek, G. A., Loeffelhard, W., and Schmetterer, G., Eds.) Kluwer Academic/Plenum, New York] have shown that the protons of water molecules that coordinate the metal can be exchanged within minutes of mixing with (2)H(2)O. Here we examine the extent and rate of deuterium exchange, using a combination of rapid freeze-quench and electron spin echo envelope modulation (ESEEM) analysis of Mn(2+)-substituted cytochrome c oxidase, which retains full activity. In the oxidized enzyme at room temperature, deuterium exchange at the Mn(2+) site occurs in less than 11 ms, which corresponds to an apparent rate constant higher than 3000 s(-1). The extent of deuterium substitution is dependent on the concentration of (2)H(2)O in the sample, indicative of rapid equilibrium, with three inner sphere (2)H(2)O exchanged per Mn(2+). This indicates that the water ligands of the Mn(2+)/Mg(2+) site, or the protons of these waters, can exchange with bulk solvent at a rate consistent with a role for this region in product release during turnover.

Animals↗
PubMed2001

Refining the ADHD phenotype using affected sibling pair families.

Attention deficit hyperactivity (ADHD) is a highly heritable behavioral disorder characterized by symptoms of inattention, hyperactivity, and/or impulsivity. Detection of susceptibility genes underlying ADHD may benefit from refinement of the ADHD phenotype into components that reflect more specific gene to behavior pathways and through the reduction of etiological heterogeneity. Using affected sibling pair (ASP) families, we are examining familial clustering of ADHD symptoms, neurocognitive task performance, and the occurrence of comorbid conditions to attempt to refine the phenotype and/or identify clinical features that may be used to stratify ADHD subjects to reduce etiological heterogeneity.

Adolescent↗
PubMed2001

Decreased anterior cingulate myo-inositol/creatine spectroscopy resonance with lithium treatment in children with bipolar disorder.

This project was designed to compare differences in brain proton spectra between children and adolescents with bipolar disorder (BPD) and gender and age-matched normal controls, and to measure changes in myo-inositol levels following lithium therapy, utilizing in vivo proton magnetic resonance spectroscopy (1H MRS). A single voxel (2x2x2 cm3) was placed in brain anterior cingulate cortex for acquisition of the 1H spectra at baseline and after acute (7 days) lithium administration in 11 children (mean age 11.4 years) diagnosed with BPD, and in 11 normal controls. Acute lithium treatment was associated with a significant reduction in the myo-inositol/creatine ratio. This decrement was also significant in lithium-responders when analyzed separate from non-responders. Compared to normal controls, BPD subjects showed a trend towards a higher myo-inositol/creatine during the manic phase. These preliminary data provide evidence that a significant reduction in anterior cingulate myo-inositol magnetic resonance may occur after lithium treatment, especially among responders. Follow-up studies involving a larger sample may allow us to confirm whether changes in myo-inositol associated with acute lithium therapy persist in long-term clinical response of patients with and without lithium compliance.

Adolescent↗
PubMed2001

Methodological issues in designing a multisite trial of risperidone in children and adolescents with autism.

OBJECTIVE: To describe the methodological challenges and decisions made in developing a multisite, controlled study of risperidone in children and adolescents with autism. METHODS: Review the design considerations for clinical trials in children with autistic disorder accompanied by severe tantrums, aggressive and/or self-injurious behaviors. These design considerations include the definition of inclusion criteria that are relevant to clinical practice and matching study design to the goal of evaluating short- and long-term effects. Additional ethical and scientific issues concern the length of trial and sample size. RESULTS: We undertook a short-term, placebo-controlled study to evaluate the efficacy and safety of risperidone in children and adolescents with autistic disorder. This trial design was followed by an extended open-label maintenance on risperidone to confirm durability of treatment effects and to monitor safety. Finally, a placebo-controlled discontinuation study tested the need for continuous treatment. CONCLUSIONS: In the absence of standard pharmacological treatment for children with autistic disorder, a placebo-controlled study remains the most appropriate method of testing efficacy and safety. The clinical relevance of this study is enhanced by the addition of an extended maintenance phase followed by a placebo discontinuation.

Adolescent↗
PubMed2000

Arachidonic acid and nonsteroidal anti-inflammatory drugs induce conformational changes in the human prostaglandin endoperoxide H2 synthase-2 (cyclooxygenase-2).

By using the technique of site-directed spin labeling combined with EPR spectroscopy, we have observed that binding of arachidonic acid and nonsteroidal anti-inflammatory drugs induces conformational changes in the human prostaglandin endoperoxide H(2) synthase enzyme (PGHS-2). Line shape broadening resulting from spin-spin coupling of nitroxide pairs introduced into the membrane-binding helices of PGHS-2 was used to calculate the inter-helical distances and changes in these distances that occur in response to binding various ligands. The inter-residue distances determined for the PGHS-2 holoenzyme using EPR were 1-7.9 A shorter than those of the crystal structure of the PGHS-2 holoenzyme. However, inter-helical distances calculated and determined by EPR for PGHS-2 complexed with arachidonic acid, flurbiprofen, and SC-58125 were in close agreement with those obtained from the cognate crystal structures. These results indicate that the structure of the solubilized PGHS-2 holoenzyme measured in solution differs from the crystal structure of PGHS-2 holoenzyme obtained by x-ray analysis. Furthermore, binding of ligands induces a conformational change in the holo-PGHS-2, converting it to a structure similar to those obtained by x-ray analysis. Proteolysis protection assays had previously provided circumstantial evidence that binding of heme and non-steroidal anti-inflammatory drugs alters the conformation of PGHS, but the present experiments are the first to directly measure such changes. The finding that arachidonate can also induce a conformational change in PGHS-2 was unexpected, and the magnitude of changes suggests this structural flexibility may be integral to the cyclooxygenase catalytic mechanism.

Animals↗
PubMed2000

Site-directed mutagenesis of 2,4-dichlorophenoxyacetic acid/alpha-ketoglutarate dioxygenase. Identification of residues involved in metallocenter formation and substrate binding.

2,4-dichlorophenoxyacetic acid (2,4-D)/alpha-ketoglutarate (alpha-KG) dioxygenase (TfdA) is an Fe(II)-dependent enzyme that catalyzes the first step in degradation of the herbicide 2,4-D. The active site structures of a small number of enzymes within the alpha-KG-dependent dioxygenase superfamily have been characterized and shown to have a similar HXDX(50-70)HX(10)RXS arrangement of residues that make up the binding sites for Fe(II) and alpha-KG. TfdA does not have obvious homology to the dioxygenases containing the above motif but is related in sequence to eight other enzymes in the superfamily that form a distinct consensus sequence (HX(D/E)X(138-207) HX(10)R/K). Variants of TfdA were created to examine the roles of putative metal-binding residues and the functions of the other seven histidines in this protein. The H167A, H200A, H213A, H245A, and H262A forms of TfdA formed inclusion bodies when overproduced in Escherichia coli DH5alpha; however, these proteins were soluble when fused to the maltose-binding protein (MBP). MBP-TfdA exhibited kinetic parameters similar to the native enzyme. The H8A and H235A variants were catalytically similar to wild-type TfdA. MBP-H213A and H216A TfdA have elevated K(m) values for 2,4-D, and the former showed a decreased k(cat), suggesting these residues may affect substrate binding or catalysis. The H113A, D115A, MBP-H167A, MBP-H200A, MBP-H245A and MBP-H262A variants of TfdA were inactive. Gel filtration analysis revealed that the latter two proteins were highly aggregated. The remaining four inactive variants were examined in their Cu(II)-substituted forms by EPR and electron spin-echo envelope modulation (ESEEM) spectroscopic methods. Changes in EPR spectra upon addition of substrates indicated that copper was present at the active site in the H113A and D115A variants. ESEEM analysis revealed that two histidines are bound equatorially to the copper in the D115A and MBP-H167A TfdA variants. The experimental data and sequence analysis lead us to conclude that His-113, Asp-115, and His-262 are likely metal ligands in TfdA and that His-213 may aid in catalysis or binding of 2,4-D.

Amino Acid Sequence↗
PubMed2000

Brain abnormalities in early-onset schizophrenia spectrum disorder observed with statistical parametric mapping of structural magnetic resonance images.

OBJECTIVE: The purpose of this study was to assess neuroanatomic abnormalities in children and adolescents with childhood-onset schizophrenia by using whole-brain voxel-based morphometric analyses. Previous volumetric studies of brain abnormalities in childhood-onset schizophrenia have revealed anomalies similar to those in subjects with adult-onset schizophrenia. Specifically, low cerebral volume, high ventricular volume, and thalamic, basal ganglia, callosal, and temporal lobe abnormalities have been observed in childhood-onset schizophrenia. Relatively few anatomical structures have been delineated and measured in this rare population, partly because of the labor involved in the slice-by-slice region definition required of conventional volumetric image analyses. METHOD: The subjects were 10 normal children and adolescents and nine children and adolescents with early-onset schizophrenia (mean age at diagnosis, 11.0 years; range, 7-16 years). The authors conducted voxel-by-voxel and volumetric statistical analyses of high-resolution structural magnetic resonance images. RESULTS: Statistical parametric maps of gray matter, white matter, and CSF differences between the groups revealed that the subjects with early-onset schizophrenia had larger ventricles, predominantly in the posterior horns of the lateral ventricles, and midcallosal, posterior cingulate, caudate, and thalamic abnormalities. Volumetric analyses of the lateral ventricles in native image data space confirmed significantly higher volume in posterior, but not anterior, regions. Randomization tests confirmed the overall statistical significance of the group differences and validity of the parametric maps. CONCLUSIONS: These findings are generally consistent with the findings of other research groups, but localization of enlarged ventricles specific to the posterior region may be a new finding in the literature on childhood-onset schizophrenia.

Adolescent↗
PubMed1999

Herbicide-degrading alpha-keto acid-dependent enzyme TfdA: metal coordination environment and mechanistic insights.

TfdA is a non-heme iron enzyme which catalyzes the first step in the oxidative degradation of the widely used herbicide (2, 4-dichlorophenoxy)acetate (2,4-D). Like other alpha-keto acid-dependent enzymes, TfdA utilizes a mononuclear Fe(II) center to activate O(2) and oxidize substrate concomitant with the oxidative decarboxylation of alpha-ketoglutarate (alpha-KG). Spectroscopic analyses of various Cu(II)-substituted and Fe(II)-reconstituted TfdA complexes via electron paramagnetic resonance (EPR), electron spin-echo envelope modulation (ESEEM), and UV-vis spectroscopies have greatly expanded our knowledge of the enzyme's active site. The metal center is coordinated to two histidine residues as indicated by the presence of a five-line pattern in the Cu(II) EPR signal, for which superhyperfine splitting is attributed to two equivalent nitrogen donor atoms from two imidazoles. Furthermore, a comparison of the ESEEM spectra obtained in H(2)O and D(2)O demonstrates that the metal maintains several solvent-accessible sites, a conclusion corroborated by the increase in multiplicity in the EPR superhyperfine splitting observed in the presence of imidazole. Addition of alpha-KG to the Cu-containing enzyme leads to displacement of an equatorial water on copper, as determined by ESEEM analysis. Subsequent addition of 2,4-D leads to the loss of a second water molecule, with retention of a third, axially bound water. In contrast to these results, in Fe(II)-reconstituted TfdA, the cosubstrate alpha-KG chelates to the metal via a C-1 carboxylate oxygen and the alpha-keto oxygen as revealed by characteristic absorption features in the optical spectrum of Fe-TfdA. This binding mode is maintained in the presence of substrate, although the addition of 2,4-D does alter the metal coordination environment, perhaps by creating an O(2)-binding site via solvent displacement. Indeed, loss of solvent to generate an open binding site upon the addition of substrate has also been suggested for the alpha-keto acid-dependent enzyme clavaminate synthase 2 [Zhou et al. (1998) J. Am. Chem. Soc. 120, 13539-13540]. Nitrosyl adducts of various Fe-TfdA complexes have also been investigated by optical and EPR spectroscopy. Of special interest is the tightly bound NO complex of Fe-TfdA.(alpha-KG).(2,4-D), which may represent an accurate model of the initial oxygen-bound species.

2,4-Dichlorophenoxyacetic Acid↗
PubMed1999

Nimodipine treatment of an adolescent with ultradian cycling bipolar affective illness.

This is a single case report of an open trial of nimodipine, a dihydropyridine-type calcium antagonist, in the treatment of a 13-year-old boy with refractory, ultradian rapid cycling, bipolar disorder type I. Prior clinical trials with calcium channel blockers in adults with ultrarapid cycling affective disorder supported an empirical trial of nimodipine for treatment of ultradian rapid cycling in this adolescent. Severity of mania and depression were rated before and after nimodipine therapy. A marked decrease in rapid, repeated, and significant mood changes was clinically observed and measured by standardized scales after 9 days of nimodipine 180 mg daily. No adverse effects were noticed. Remission persisted with continued treatment at 36-month follow-up. Medication response was partially attributed to adjunctive therapy with levothyroxine. Implications of treatment benefit are discussed in the context of novel pharmacotherapies for refractory bipolar disorder. These findings are preliminary and do not provide sufficient basis to recommend nimodipine as the treatment of choice in adolescents with ultradian cycling bipolar disorder, but suggest that controlled studies may be indicated.

Activity Cycles↗
PubMed1998

Manganese and tyrosyl radical function in photosynthetic oxygen evolution.

Photosystem II catalyzes the photosynthetic oxidation of water to O2. The structural and functional basis for this remarkable process is emerging. The catalytic site contains a tetramanganese cluster, calcium, chloride and a redox-active tyrosine organized so as to promote electroneutral hydrogen atom abstraction from manganese-bound substrate water by the tyrosyl radical. Recent work is assessed within the framework of this model for the water oxidizing process.

Binding Sites↗
PubMed1997

Tyrosyl radicals in enzyme catalysis: some properties and a focus on photosynthetic water oxidation.

Enzymes that require a redox-active amino acid for catalysis or function have emerged as a distinct class of proteins. For the tyrosine-based radical enzymes, we show that the spin-density distribution in the radical follows an odd alternate pattern that is invariant to within 10% across the class. General properties of the radical enzymes are summarized from which we conclude that their essential role in catalysis is to initiate substrate metabolism by hydrogen-atom abstraction. These ideas are extended to the YZ and YD tyrosines in Photosystem II and a radical-based hydrogen-atom abstraction model for water oxidation is discussed. Differences in rates of oxidation of YZ and YD by the reaction-center chlorophyll, P680+, under various conditions, are considered and rationalized on the basis of changes in reorganization energy induced by the local protein structure and by the presence or absence of the (Mn)4 cluster that binds substrate water.

Free Radicals↗
PubMed1996

Identification of histidine as an axial ligand to P700+.

The primary electron donor in photosystem I (PSI), P700, is thought to be a dimeric Chl a species. Neither the electronic nor geometric structure of the cation radical is clearly understood. Magnetic resonance studies have indicated that the unpaired electron in P700+ is delocalized asymmetrically over the Chl dimer; however, the axial ligand to the central Mg2+ is not known. The recent development of a histidine tolerant mutant of Synechocystis PCC 6803 has allowed us to use a combination of isotopic labeling and electron nuclear double resonance (ENDOR) spectroscopy to show the first definitive spectroscopic evidence of a histidine ligand to P700+. Peaks split symmetrically about the 15N Larmor frequency corresponding to an isotropic hyperfine coupling of 0.64 MHz were observed in the ENDOR spectra from P700+ globally labeled with 15N and specifically labeled with [15N]histidine. These peaks disappeared in "reverse" labeled samples in which all nitrogens are 15N except those of histidine, which contains natural abundance 14N. The dipolar contribution to the hyperfine coupling was determined by using electron spin echo envelope modulation spectroscopy (ESEEM). Numerical simulations of the ESEEM data suggest that the coupling is primarily isotropic and that the histidine is directly coordinated to the central Mg2+ of P700+. Taken together, these data are supportive of a model of P700+ in which the excited state molecular orbital makes a significant contribution to the electronic structure of the radical. Moreover, the methodology developed in this work can be extended to examine the magnetic properties of axial ligands in a variety of biologically relevant porphyrin/chlorin systems.

Binding Sites↗
PubMed1995

A chemical modification of cytochrome-c lysines leading to changes in heme iron ligation.

Although 13 lysines of horse cytochrome c are invariant, and three more are extremely conserved, the modification of their side-chain epsilon-amino groups by beta-thiopropionylation caused important changes in protein properties for only three of them; lysines 72,73 and 79. Optical spectroscopy, electron and nuclear paramagnetic resonance, electron spin echo envelope modulation, and molecular weight studies, as well as the unique features of their reaction with cytochrome-c oxidase, indicate that in the oxidized state the modification of these lysines resulted in equilibria between two different states of iron ligation: the native state, in which the metal is coordinated by the methionine-80 sulfur, and a new state in which this ligand is displaced by the sulfhydryl groups of the elongated side chains. The reduction potentials of the TP Lys-72 and the TP Lys-79 derivatives were 201 and 196 millivolt, respectively, indicating that the equilibria favored the sulfhydryl ligated state by 1.5 and 1.7 kcal/mol, respectively. In the ferric state, the protein modified at lysine 72 remained stable as a monomer, but that modified at lysine 73 dimerized rapidly through disulfide bond formation, while the TP Lys-79 cytochrome c dimerized with a half-time of approx. 3 h, both recovering the native-like iron ligation. By contrast, in the ferrous state the monomeric state and the native ligation were preserved in all cases, indicating that the affinity of the cytochrome-c ferrous iron for the methionine-80 sulfur is particularly strong. The dimerized derivatives lost most, but not all, of the capability of the native protein for electron transfer from ascorbate-TMPD to cytochrome-c oxidase.

Animals↗
PubMed1995

A continuous wave and pulsed EPR characterization of the Mn2+ binding site in Rhodobacter sphaeroides cytochrome c oxidase.

The ligation environment of the tightly bound Mn2+ in cytochrome c oxidase from Rhodobacter sphaeroides has been characterized by electron paramagnetic resonance (EPR) and electron spin echo envelope modulation (ESEEM). The EPR data show that the Mn2+ is six-coordinate and located in a highly symmetric binding site. Analyses of X- and Q-band EPR spectra show that the zero field splitting parameter D is 115 +/- 25 G (0.0107 +/- 0.0023 cm-1) in the fully oxidized enzyme and 125 +/- 15 G (0.0117 +/- 0.0014 cm-1) in the fully reduced enzyme. For both redox forms of the enzyme the value of E is < or = 25 G (0.0023 cm-1). By comparison with crystal structures of Mn2+ binding proteins, the structural changes at the Mn2+ binding site upon redox state change of the enzyme are estimated to be < or = 0.2 A in ligand bond lengths and < or = 10 degrees in bond angle. This analysis indicates that little modification occurs at the Mn2+ site upon redox change at the other metal centers. Considering the proximity of the Mn2+ site to heme a and heme a3-CuB [Hosler, J. P., Espe, M. P., Zhen, Y., Babcock, G. T., & Ferguson-Miller, S. (1995) Biochemistry 34, 7586-7592], we interpret these results to imply also that there is no large protein conformational change near the heme a and heme a3-CuB sites upon a change in their redox states. Multifrequency 3-pulse ESEEM results provide direct evidence for a nitrogen ligand to the Mn2+, which is assigned to a histidine by comparison with ESEEM studies of Mn(2+)-bound lectins [McCracken, J., Peisach, J., Bhattacharyya, L., & Brewer, F. (1991) Biochemistry 30, 4486-4491] and specifically to His-411 in subunit 1 on the basis of mutagenesis studies (Hosler et al., 1995). From these results a partial model of the Mn2+ binding site has been constructed.

Binding Sites↗
PubMed1995

Autism, affective disorders, and social phobia.

The purpose of this study is to test the hypothesis that major affective and/or anxiety disorders are increased among relatives of autistic probands compared with controls. Among 36 families with an autistic child, 23 (64%) have a first degree relative diagnosed with major depressive disorder and 14 (39%) have a first degree relative diagnosed with social phobia. These rates are significantly greater than the 19% and 5%, respectively, found among 21 families with a child having a genetic condition, tuberous sclerosis complex, or a seizure disorder but no autism. The frequency of major depression among the 96 first degree relatives of autistic probands is 37.5% compared with 11.1% found among 45 relatives of control probands. The frequency of social phobia, 20.2%, is approximately 10 times more common than that found among the relatives of the control probands (2.4%). Elevated rates of both major depression and social phobia are found among parents and siblings in the families with an autistic child. Furthermore, 64% of parents affected with a major depression had the onset of the first depressive episode prior to the birth of the autistic child and all parents with social phobia had the onset of condition prior to the birth of the autistic child. Family patterns differ depending on the intellectual level of the autistic child; specifically, social phobia is significantly greater among the first degree relatives of non-retarded autistic probands than among relatives of individuals with autism and comorbid mental retardation.(ABSTRACT TRUNCATED AT 250 WORDS)

Autistic Disorder↗
PubMed1995

Cu(II) coordination in arthropod and mollusk green half-methemocyanins analyzed by electron spin-echo envelope modulation spectroscopy.

Hemocyanin (Hc) is a dinuclear copper protein that binds oxygen reversibly. The structure of the Cu(II) site in a derivative of hemocyanin known as green half-met (GHM) has been analyzed using the pulsed EPR technique of electron spin-echo envelope modulation (ESEEM) spectroscopy. The derivative, prepared by treating the native protein with nitrite at low pH, contains a mixed-valent binuclear copper center. It was shown through chemical assays and the ligand exchange reaction products identified by EPR spectroscopy to contain a nitrite ligand bound to Cu(II). The ESEEM spectra of green half-methemocyanins from mollusks and arthropods indicated that three imidazole ligands are coordinated to Cu(II). Therefore, a tetragonal N3O ligand structure (O is an oxygen of nitrite) is proposed. For GHM Hc from the mollusks Octopus vulgaris and Rapana thomasiana, the isotropic nitrogen nuclear hyperfine coupling constant, aiso, for the N delta (or remote) nitrogen of two imidazoles was approximately 1.4 MHz, while for the third, aiso congruent to 2.2 MHz. The difference between the two weaker nitrogens and the single, more strongly coupled nitrogen was smaller by 0.2 MHz in the GHM Hcs from the arthropods Carcinus maenas, Homarus americanus and Panulirus interruptus. The nitrogen nuclear quadrupole coupling constants and asymmetry parameters, e2Qq and eta, for the N delta nitrogens in nearly all cases were near 1.4 MHz and 0.8, respectively, although Rapana thomasiana GHM Hc exhibited a reduction in eta that may indicate weaker hydrogen bonding in the active site of this protein. The g and ACu (copper nuclear hyperfine coupling) values for the derivatives, and the finding of three similar nuclear hyperfine coupling constants for the N delta sites of imidazole ligands, when considered with the orientation-specific information obtained using angle-selection methods for simulation of ESEEM spectra, suggest a distorted tetragonal Cu(II) structure in which three imidazoles and a nitrite ligand are bound near the equatorial plane. The finding that the two molluscan GHM Hcs exhibit differences associated with the remote nitrogen of imidazoles bound to Cu(II) may be related to a structural variability in the active sites of these proteins not found in the arthropodan GHM Hcs examined.

Animals↗