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Biomedical subjects

J Carey

Publications and source records attributed to J Carey.

At least 73 records · Page 4Linked to original sources
PubMed1993

Cocrystals of Escherichia coli trp repressor bound to an alternative operator DNA sequence.

Cocrystals of a 2:1 complex of trp repressor dimers with a DNA duplex containing a single, central operator half-site sequence are described. Crystals with different morphologies grew under diverse crystallization conditions within days to weeks by hanging-drop vapor-diffusion. Twinned rods split along their longitudinal cleft produce single crystals with space group C2 and unit cell dimensions a = 112.34 A, b = 90.16 A, c = 58.65 A and beta = 113.92 degrees. The crystals diffract to 2.4 A and are thus suitable for structural analysis by X-ray diffraction. Several heavy-atom derivative cocrystals have been obtained with iodouridine-substituted DNAs.

Bacterial Proteins↗
PubMed1993

Tandem binding in crystals of a trp repressor/operator half-site complex.

The crystal structure of trp repressor tandemly bound in a 2:1 complex to a 16-base-pair palindromic DNA containing a central trp operator half-site has been determined and refined to 2.4 A resolution. Despite dramatically different DNA sequence contexts and crystallization conditions, the protein/DNA interface is essentially identical to that seen in the original trp repressor/operator complex structure. Water-mediated sequence recognition by trp repressor is likely to be related to the unusual end-on approach of the recognition helix (E), which allows sharing of the major groove by tandem dimers. The tandem complex model accounts for the mutational sensitivity of all trp operator base pairs. The structure also provides the first detailed view of the tandem interaction, revealing a key role for the amino-terminal arms.

Bacterial Proteins↗
PubMed1993

Prenatal detection of de novo paracentric inversion 46, XX inv (14) (q22q32.1) in a normal child: report and review of the literature.

We present prenatal diagnosis and follow-up examination of an individual with a de novo paracentric inversion of the long are of chromosome 14. A literature search documented 19 other cases of paracentric inversion of 14q. The outcome of each of these cases is specified together with that of this current case. Four of the 20 cases, all XY, manifested significant abnormalities with mental retardation and microcephaly present in 3 of the 4 cases; 15% (2/13) of familial cases had abnormalities and 40% (2/5) of de novo cases had abnormalities.

Adult↗
PubMed1993

A noncovalent peptide complex as a model for an early folding intermediate of cytochrome c.

Horse heart cytochrome c is one of a small number of proteins for which the folding pathway has been elucidated in structural detail by pulsed hydrogen exchange and NMR. Those studies indicated that a partially folded intermediate with interacting N- and C-terminal helices is formed at an early stage of folding when most of the chain is still disordered. This report describes a peptide model for this early intermediate, consisting of a noncovalent complex between a heme-containing N-terminal fragment (residues 1-38) and a synthetic peptide corresponding to the C-terminal helix (residues 87-104). Far-UV circular dichroism and proton NMR indicate that the isolated peptides are largely disordered, but when combined, they form a flexible, yet tightly bound complex with enhanced helical structure. These results emphasize the importance of interactions between marginally stable elements of secondary structure in forming tertiary subdomains in protein folding.

Amino Acid Sequence↗
PubMed1993

Thermodynamics of ligand binding to trp repressor.

The thermodynamics of L-tryptophan and operator DNA binding to the tryptophan repressor of Escherichia coli were analyzed by titration microcalorimetry and van't Hoff analysis of footprinting titrations, respectively. At 25 degrees C in 10 mM sodium phosphate, pH 7.6, and 0.1 M NaCl, the binding of L-tryptophan to the repressor is characterized by values of delta G degrees = -6.04, delta H degree = -14.7, and T delta S degree = -8.67 kcal/mol. The temperature dependence of delta H degree yields delta Cp degree = -0.46 +/- 0.08 kcal/(mol.K) per dimer. The binding is noncooperative at all temperatures studied. At 23 degrees C in 2.5 mM sodium phosphate, pH 7.6, and 25 mM NaCl, the binding of operator DNA to the repressor is characterized by values of delta G degree = -13.3 kcal/mol, delta H degree = -1.55 kcal/mol, T delta S degree = 11.8 kcal/mol, and delta Cp degree = -0.54 +/- 0.10 kcal/(mol.K). Changes in water-accessible surface areas upon binding of L-tryptophan or DNA were calculated from X-ray crystal structures. The experimentally observed delta Cp degree values were compared with delta Cp degree values calculated according to several methods based on various proposed relationships between surface area changes and heat capacity changes. Regardless of which method is used, we find poor agreement between the calorimetric results for L-tryptophan binding and the surface areas calculated from X-ray data; the direction of the discrepancy is that the X-ray data underestimate the value of delta Cp degree.(ABSTRACT TRUNCATED AT 250 WORDS)

Bacterial Proteins↗
PubMed1993

Reversal of caldesmon binding to myosin with calcium-calmodulin or by phosphorylating caldesmon.

Caldesmon, an actin-binding protein from smooth muscle and non-muscle cells, has previously been shown to bind stoichiometrically to smooth muscle myosin in an ATP-dependent manner. We now show quantitatively the effects of Ca(2+)-calmodulin and phosphorylation on the binding of caldesmon to myosin. Ca(2+)-calmodulin reduces the binding of caldesmon to myosin with the same effectiveness as it does the binding of caldesmon to actin. However, Ca(2+)-calmodulin is ineffective in antagonizing the binding of the purified myosin-binding region of caldesmon to myosin. These and other results suggest that Ca(2+)-calmodulin binding to the COOH-terminal region of caldesmon is responsible for reversal of binding to myosin. Phosphorylation of the NH2-terminal region of caldesmon by the co-purifying kinase, calmodulin-dependent protein kinase II, weakens but does not eliminate the binding of caldesmon to smooth muscle myosin. Finally, phosphorylation of smooth muscle myosin by smooth muscle myosin light chain kinase has no effect on the binding of caldesmon to myosin. Since Ca(2+)-calmodulin and phosphorylation of caldesmon weaken the binding of caldesmon to both actin and myosin, these events may be coordinately regulated.

Adenosine Triphosphate↗
PubMed1993

Analysis of trp repressor-DNA interactions using gel electrophoresis.

Quantitative analysis of the DNA-binding equilibria of E. coli trp repressor by gel electrophoresis led to reevaluation of our understanding of this complex system. In this review, the data leading to controversy about the trp system are discussed, and our current understanding is presented.

Bacterial Proteins↗
PubMed1993

Comparison of VNTR allele frequencies and inclusion probabilities over six populations.

There is considerable debate about the methodologies used to estimate VNTR (Variable Number of Tandem Repeats) multi-locus genotype frequencies or odds of inclusion in forensic cases. To compare two of the methods in use, allele frequency distributions among six populations were compared and the effect of population heterogeneity on VNTR multi-locus genotype frequency estimation was examined. Genotype frequencies estimated from single population data were one or two orders of magnitude smaller than those estimated by picking the highest allele frequency in a group of subpopulations to estimate genotype frequencies using a ceiling principle. The average change does not appear to be very sensitive to the set of subpopulations used; four locus frequencies still give inclusion odds of one in a million or less. We think that use of the ceiling principle solves both the statistical problem engendered by subpopulation heterogeneity and the legal problem of assuming that the prepetrator and suspect belong to the same subpopulation. The counterintuitive fact of human genetic polymorphism is that it is easier to identify an individual than it is to identify the subpopulation, ethnic group or race to which that individual belongs.

Africa↗
PubMed1992

Recombinant human erythropoietin in the treatment of anemia associated with human immunodeficiency virus (HIV) infection and zidovudine therapy. Overview of four clinical trials.

OBJECTIVE: To assess the effect of recombinant human erythropoietin (r-HuEPO) on anemia in patients with the acquired immunodeficiency syndrome (AIDS) who are receiving zidovudine therapy. DESIGN: Combined analysis of four 12-week, randomized, double-blind, controlled clinical trials. SETTING: Multiple centers in the United States. PATIENTS: Two hundred and ninety-seven anemic (hematocrit < 30%) patients with AIDS who were receiving zidovudine therapy. Of the 297 patients, 255 were evaluable for efficacy, but all patients were included in analysis of safety. INTERVENTION: Patients were randomly assigned to receive either r-HuEPO (100 to 200 U/kg body weight) or placebo, intravenously or subcutaneously, three times per week for up to 12 weeks. MEASUREMENTS: Changes in mean hematocrit, transfusion requirement, and quality of life. RESULTS: Sixty-nine percent of patients had endogenous serum erythropoietin levels less than or equal to 500 IU/L, and 31% had erythropoietin levels greater than 500 IU/L. In patients with low erythropoietin levels (< or equal to 500 IU/l), r-HuEPO therapy decreased the mean number of units of blood transfused per patient when compared with placebo (3.2 units and 5.3 units, respectively; P = 0.003) and increased the mean hematocrit from the baseline level (4.6 percentage points and 0.5 percentage points, respectively; P <0.001). Overall quality of life improved in patients on r-HuEPO therapy (P = 0.13). Patients with erythropoietin levels greater than 500 IU/L showed no benefit from r-HuEPO in any outcome variable. Placebo and r-HuEPO recipients did not differ in the incidence of adverse effects or opportunistic infections. CONCLUSION: Therapy with r-HuEPO can increase the mean hematocrit and decrease the mean transfusion requirement in anemic patients with AIDS who are receiving zidovudine and have endogenous low erythropoietin levels (< or equal to 500 IU/L). Such therapy is of no apparent benefit in patients whose endogenous erythropoietin levels are higher than 500 IU/L.

Acquired Immunodeficiency Syndrome↗
PubMed1992

Binding of the arginine repressor of Escherichia coli K12 to its operator sites.

In the arginine regulon of Escherichia coli K12 each of the eight operator sites consists of two 18-base-pair-long palindromic sequences called ARG boxes. In the operator sites for the structural genes of the regulon the two ARG boxes are separated by three base-pairs, in the regulatory gene argR they are separated by two base-pairs. The hexameric arginine repressor, the product of argR, binds to the two ARG boxes in an operator in the presence of L-arginine. From the results of various kinds of in vitro footprinting experiments with the ARG boxes of argF and argR (DNase I protection, hydroxyl radical, ethylation and methylation interference, methylation protection) it can be concluded that: (1) the repressor binds simultaneously to two adjacent ARG boxes; (2) that it binds on one face of the double helix; and (3) that it forms contacts with the major and minor grooves of each ARG box, but not with the central three base-pairs. The repressor can bind also to a single ARG box, but its affinity is about 100-fold lower than for two ARG boxes. From gel retardation experiments with 3H-labeled repressor and 32P-labeled argF operator DNA, it is concluded that the retarded DNA-protein complex contains no more than one repressor molecule per operator site and that most likely one hexamer binds to two ARG boxes. The bound repressor was shown to induce bending of argF operator DNA. The bending angle calculated from the results of gel retardation experiments is about 70 degrees and the bending center was located within the region encompassing the ARG boxes. The main features that distinguish the arginine repressor from other repressors studied in E. coli are its hexameric nature and the simultaneous binding of one hexameric molecule to two palindromic ARG boxes that are close to each other.

Arginine↗
PubMed1992

Anatomical organization of the brainstem octavolateralis area of the oyster toadfish, Opsanus tau.

Anatomical studies were undertaken to analyze the brainstem organization of the auditory, vestibular, and lateral line nuclei in a teleost, the oyster toadfish, Opsanus tau. Neuronal cytoarchitectonics and horseradish peroxidase label of cranial nerves were utilized to delineate the borders of the five octavus and two lateralis brainstem nuclei. Each of the eight octavolateralis nerves were labeled individually to compare and contrast their central projections. Projections of the three semicircular canals were found to be largely overlapping. Terminal fields were observed within the eminentia granularis and in each of the octavus nuclei. The nucleus anterior octavus was reciprocally innervated by the semicircular canals and the saccule. The canals terminated heavily in the ventral portions of the anterior octavus, whereas the saccule terminated extensively in the dorsal nuclear portions. The saccule also distributed terminals throughout the octavus cell column, including a light terminal field within the dorsal, medial, and anterior portions of the descending octavus nucleus, a region densely innervated by this end-organ in other species. These results suggest that the anterior octavus nucleus may have a dual function. The dorsal portions may be an auditory relay nucleus, whereas the ventral portions may subserve vestibular function. Utriclar and lagenar afferents also terminated throughout the octavus cell column. Afferents of the anterior and posterior lateral lines ended within the eminentia granularis and the lateral line nuclei. Semicircular canal afferents and lateral line afferents appeared completely segregated within the eminentia. The above results are useful as an aid in the understanding of an ongoing, comprehensive functional analysis of auditory and vestibular mechanisms in toadfish and complement previous work on the efferent vestibular and sound-producing motor systems. Examination of toadfish contributes to a more general and complete overview of the octavolateralis area of teleosts and the eventual identification of primitive and derived patterns of octaval organization. Additionally, this work may permit the further demonstration of species-typical characters that may indicate adaptations to particular behavioral repertoires.

Animals↗
PubMed1992

Ordered self-assembly of polypeptide fragments to form nativelike dimeric trp repressor.

Subdomain-size proteolytic fragments of Escherichia coli trp repressor have been produced that assemble in defined order to regenerate fully native dimers. By characterization of the secondary and tertiary structures of isolated and recombined fragments, the structure of assembly intermediates can be correlated with the kinetic folding pathway of the intact repressor deduced from spectroscopic measurement of folding rates. The nativelike structure of these intermediates provides further evidence that protein folding pathways reflect the stabilities of secondary structural units and assemblies found in the native state. The proteolytic method should be generally useful in adding structural detail to spectroscopically determined folding mechanisms.

Amino Acid Sequence↗
PubMed1992

What makes information important and its influence on learning in self-directed study.

This study investigated what makes text-based information important and what is its influence on learning. College students studied an expository text following their own self-directed study procedures. Data from Exp. 1 (n = 60) and Exp. 2 (n = 40) showed that information was considered important if it (a) began and ended text, (b) began a paragraph, (c) appeared at somewhat equal intervals in text, and (d) contained enumerations of concepts. Subjects recalled information that (a) began the text and (b) contained examples of enumerations. It appeared that text-based variables established what was important and this influenced learning. Implications for the selective attention hypothesis and its volume-of-attention corollary, the cycles hypothesis, and the slot-filling hypothesis are discussed.

Achievement↗
PubMed1992

What are trusts allowed to do?

NHS trusts may not be a feature of the health services for ever, but current legislation will be applicable for some time to come. Legal aspects of trust status are described in two features. In the first, Anita Morrish identifies what trusts are legally empowered to do and those areas where their powers, if they exist, are less clearly defined.

Hospital Restructuring↗
PubMed1992

Treatment of cryptosporidiosis with paromomycin. A report of five cases.

Cryptosporidiosis continues to be one of the most devastating complications of the acquired immunodeficiency syndrome, causing severe, chronic diarrhea that is largely refractory to treatment. More than 60 drugs have been tried in the treatment of cryptosporidiosis, none of which have been consistently successful. We describe the successful treatment of cryptosporidiosis in five patients with acquired immunodeficiency syndrome with oral paromomycin at a dose of 1500 to 2000 mg/d. All five patients had resolution of symptoms and normalization of bowel movements, although one patient later relapsed while receiving paromomycin. Three of five patients cleared Cryptosporidium from the stool. Paromomycin is a promising therapy for cryptosporidiosis in acquired immunodeficiency syndrome and further prospective clinical trials are warranted.

AIDS-Related Opportunistic Infections↗
PubMed1991

How does trp repressor bind to its operator?

Three explanations have been advanced to account for the unexpected absence of direct hydrogen bonds and presence of a buried water layer in the co-crystal-line complex of Escherichia coli trp repressor with DNA. We present results of physical and biochemical measurements that address the testable predictions of each model. We find that the DNA oligomer used for co-crystallization binds to the repressor with high affinity and specificity, and 1:1 stoichiometry, consistent with other evidence that this sequence represents the true operator target for a single repressor dimer. A proposed alternative DNA sequence binds weaker and with higher stoichiometry, consistent with a cooperative binding mode. The operator DNA in solution has a B-form helical structure in the presence and absence of repressor. Affinity of repressor for operator is altered under the conditions used for cocrystal growth.

Bacterial Proteins↗
PubMed1991

Clindamycin and primaquine as primary treatment for mild and moderately severe Pneumocystis carinii pneumonia in patients with AIDS.

The combination of clindamycin and primaquine was investigated as primary treatment for mild or moderately severe Pneumocystis carinii pneumonia (alveolar-arterial oxygen gradient less than 40 torr) in 36 AIDS patients. All patients were treated with primaquine 30 mg (base) orally for 21 days. The first 22 patients were treated with clindamycin 900 mg i.v. q8h for the first 10 days, followed by clindamycin 450 mg q6h orally for 11 days. Twenty of these 22 patients (91%) showed a marked clinical response by day 7. Four patients were later withdrawn from the study due to toxicity. An additional 14 patients were treated with an entirely oral regimen of clindamycin and primaquine. Thirteen of the 14 patients (93%) showed a marked therapeutic response and 12 (86%) successfully completed treatment. The combination of clindamycin and primaquine appears to be an effective and well-tolerated treatment for mild to moderately severe Pneumocystis carinii pneumonia in AIDS patients.

Acquired Immunodeficiency Syndrome↗