Biomedical subjects

M S Johnson

Publications and source records attributed to M S Johnson.

At least 163 records · Page 9Linked to original sources

PubMed1994

The recognition of protein structure and function from sequence: adding value to genome data.

The explosion of DNA sequence data from genome projects presents many challenges. For instance, we must extend our current knowledge of protein structure and function so that it can be applied to these new sequences. The derivation of rules for the relationships between sequence and structure allow us to recognize a common fold by the use of tertiary templates. New techniques enable us to begin to meet the challenge of rule-based modelling of distantly related proteins. This paper describes an integrated and knowledge-based approach to the prediction of protein structure and function which can maximize the value of sequence information.

Amino Acid Sequence↗

PubMed1994

Residue-residue contact substitution probabilities derived from aligned three-dimensional structures and the identification of common folds.

We report the derivation of scores that are based on the analysis of residue-residue contact matrices from 443 3-dimensional structures aligned structurally as 96 families, which can be used to evaluate sequence-structure matches. Residue-residue contacts and the more than 3 x 10(6) amino acid substitutions that take place between pairs of these contacts at aligned positions within each family of structures have been tabulated and segregated according to the solvent accessibility of the residues involved. Contact maps within a family of structures are shown to be highly conserved (approximately 75%) even when the sequence identity is approaching 10%. In a comparison involving a globin structure and the search of a sequence databank (> 21,000 sequences), the contact probability scores are shown to provide a very powerful secondary screen for the top scoring sequence-structure matches, where between 69% and 84% of the unrelated matches are eliminated. The search of an aligned set of 2 globins against a sequence databank and the subsequent residue contact-based evaluation of matches locates all 618 globin sequences before the first non-globin match. From a single bacterial serine proteinase structure, the structural template approach coupled with residue-residue contact substitution data lead to the detection of the mammalian serine proteinase family among the top matches in the search of a sequence databank.

Amino Acid Sequence↗

PubMed1994

Fluoride accumulation and toxicity in wild small mammals.

Populations of two species of small mammal, the field vole (Microtus agrestis L.) and the bank vole (Clethrionomys glareolus L.), inhabiting grasslands contaminated by industrial sources of fluoride were examined for fluoride concentrations in skeletal tissue and for morphological changes in the teeth. Concentrations of fluoride in teeth and bones were higher for C. glareolus than for M. agrestis at the chemical works and smelter sites. Severe dental lesions were recorded on the incisor and molar teeth of both species at the chemical works and smelter sites, with less marked damage at the mine tailings dam. This is attributed to inter-site differences in fluoride speciation and the consequent effects on the availability of fluoride in the diet for bioassimilation.

Journal Article↗

PubMed1994

Protein structure comparisons using a combination of a genetic algorithm, dynamic programming and least-squares minimization.

We introduce a completely automatic and objective procedure for the comparison of protein structures. A genetic algorithm is used to search for a near optimal solution of the rigid-body superposition of two whole protein structures. The specification of an initial set of equivalences is not required. Topological equivalences in the final structural alignment are defined by a conventional dynamic programming routine, which is commonly used to compare protein sequences. A least-squares fitting algorithm is then used to optimize the fit between the final set of equivalences. We have applied our method to the comparison of ribonucleic acid structures, as well as protein structures. The structural alignments are generally consistent with those previously published. In fact, on most occasions our method defines at least the same number of topological equivalences as other procedures, but always with a lower r.m.s. distance between them.

Algorithms↗

PubMed1994

Intrahepatic stones. The transhepatic team approach.

OBJECTIVE: The authors reviewed the combined interventional radiologic and surgical management of 54 patients with intrahepatic stones at the Johns Hopkins Hospital. The team approach used large-bore transhepatic stents to access the intrahepatic ducts until they were stone free. SUMMARY BACKGROUND DATA: Intrahepatic stones are uncommon in western countries. As a result, few American institutions have had much experience, and multiple management algorithms have been suggested. Nonoperative, operative, and combination surgical and nonoperative approaches have been advocated. At Johns Hopkins, combined surgical and percutaneous management has been used for 18 years. METHODS: This team approach includes (1) percutaneous placement of transhepatic access catheters, (2) surgery for underlying biliary disease and stone removal, and, when necessary (3) postoperative percutaneous choledochoscopy and stone removal through the transhepatic stents. RESULTS: The median age of the 54 patients was 50 years, and 32 were men. Biliary disease included 27 benign strictures, 7 sclerosing cholangitis, 5 choledochal cysts, 5 parasitic infections, 5 choledocholithiasis, and 5 biliary tumors. Fourteen patients (26%) were treated exclusively with percutaneous techniques. Forty patients (74%) had surgery, including 36 Roux-en-Y hepatico- or choledochojejunostomies with large-bore transhepatic stents. Eighteen of these 40 patients (45%) with multiple intrahepatic stones, strictures, or both required additional procedures after operation. No hospital deaths occurred after any of the percutaneous or surgical procedures. With a mean follow-up of 60 months, 94% of patients were stone free, 87% of patients were symptom free, and 73% have had their transhepatic stents removed. CONCLUSIONS: A combined radiologic and surgical approach with transhepatic stents is a safe and effective method for managing intrahepatic stones.

Adult↗

PubMed1994

The persistent computed tomography nephrogram: its significance in the diagnosis of contrast-associated nephrotoxicity.

A study was undertaken to find out if there is a quantitative relationship between the development of contrast-associated nephrotoxicity (CAN) and the mean renal cortical attenuation (RCA) as seen on computed tomography (CT) examination of the kidneys 24 hours after injection of contrast medium. 96 patients undergoing aortography were selected, who were considered "high risk" either because of the presence of a baseline serum creatinine (Cr) of 123.76 mumol l-1 (1.4 mg dl-1) or higher, or who were 73 or older with or without elevated creatinine levels. A CT study of the kidneys at approximately 24 h was correlated with baseline serum creatinine as well as 24 h Cr (48-72 h Cr when available). Medical history and medications were recorded. Eight of 96 patients had RCA of 108-236 HU at 24 h CT and all developed significant CAN of varying degree; 3/96 had RCA ranging from 48 to 67 HU and had transient CAN; two other patients with RCA of 67-90 HU developed CAN; and there was increased incidence of CAN in patients: (a) with pre-existing renal insufficiency; (b) with diabetes with renal insufficiency; (c) on diuretics; (d) who were older; and (e) had a unilateral atrophic kidney. It was concluded that a 24 h CT study evaluating the RCA had better predictive value for the development of CAN than a 24 h creatinine level.

Age Factors↗

PubMed1994

Activation of MAP kinase associated with the priming effect of LHRH.

A MAP kinase activity assay was developed to determine whether the LHRH receptor could activate this enzyme (particularly during LHRH priming). In anterior pituitary tissue from prooestrous rats LHRH caused concentration-dependent activation of MAP kinase after 5-10 min and continued for up to 60 min of incubation. The magnitude of this response correlated with that of LHRH priming on various days of the oestrous cycle but not with the magnitude of 1st hour (unprimed) LHRH-induced LH release. The response to LHRH was mimicked by a phorbol ester but not by ionomycin and was blocked with high potency by GF 109203X but not by H7 (in a similar manner to the PKC species that mediates LHRH priming). Neither the tyrosine kinase inhibitor lavendustin A nor the protein synthesis inhibitor cycloheximide blocked LHRH-induced MAP kinase activation. The possible functional significance of MAP kinase activation in gonadotrophs is considered with respect to LHRH priming.

Animals↗

PubMed1994

Evidence for a role of phospholipase A2 in the mechanism of LHRH priming in rat anterior pituitary tissue.

The phospholipase A2 (PLA2) inhibitors, quinacrine, p-bromophenacyl bromide, ONO-RS-082, aristolochic acid and chloracysine blocked the priming effect of LHRH, but not acute LHRH-induced gonadotrophin release measured in anterior pituitary pieces in pro-oestrous rats in vitro. These results suggest that the intracellular mechanisms underlying LHRH priming are distinct from those which mediate LH release in the present circumstances in that they involve PLA2. Furthermore, neither LHRH-induced LH release from preprimed tissue nor Ca(2+)-induced LH release were attenuated by quinacrine, indicating that this inhibitor does not interfere with the general Ca(2+)-dependent secretory apparatus of the gonadotroph and that the critical period for its action is in the induction of priming. LHRH induced the release of [3H]arachidonic acid ([3H]AA) from [3H]AA-prelabelled anterior pituitary tissue from pro-oestrous rats; a response which was sensitive to inhibitors of PLA2, of protein kinase C (PKC) and of protein synthesis. Activation of PKC also resulted in [3H]AA release which was inhibited with exactly the same pharmacological profile as the response to LHRH. Both gonadotrophin secretion and [3H]AA release responses to LHRH and to phorbol ester varied in parallel during the oestrous cycle and in ovariectomized/oestradiol-17 beta-replaced animals, as did their sensitivity to quinacrine and the protein synthesis inhibitor cycloheximide. These results indicate that LHRH priming is dependent on a hormonally regulated cascade involving a distinct form of PKC acting through a protein synthesis-dependent step to release AA by means of PLA2 activity. The priming effect was mimicked (at least in part) by conditioning preincubation with AA, confirming the functional relevance of this signalling cascade. Results using standard inhibitors of lipoxygenase/epoxygenase pathways were equivocal as to whether these pathways were critically involved, whilst cyclo-oxygenase inhibitors were completely without effect. The steps downstream from AA (and its possible metabolites) by which stimulus-secretion coupling is up-regulated in priming remain to be clarified.

Acetophenones↗

PubMed1994

Knowledge-based protein modeling.

Knowledge, both from the three-dimensional structures of homologous proteins and from the general analysis of protein structure, is of value in modeling a protein of known sequence but unknown structure. While many models are still constructed at least in part by manual methods on graphics devices, automated procedures have come into greater use. These procedures include those that assemble fragments of structure from other known structures and those that derive coordinates for the model from the satisfaction of restraints placed on atomic positions.

Amino Acid Sequence↗

PubMed1994

A systematic review of the Journal of the National Medical Association 1992.

An analysis of the articles published in 1992 in the Journal of the National Medical Association (JNMA) was performed to assess the content, statistical analysis, and the profile of its contributors. Seventy-six articles were reviewed. The majority of the articles focused on biomedical aspects of health; however, a significant proportion of the remaining articles contained information on the psychosocial aspects of health as it relates to people of African descent in the United States, Africa, and the Caribbean. Most of the literature was derived from physicians in university settings, with the contributions from traditionally black medical schools constituting nearly 30% of the articles. Analysis of data from contingency tables was the most common research method used. This study suggests that the JNMA is contributing to the mission of the National Medical Association. Reasons why authors who perform research with more rigorous scientific method do not publish in JNMA should be explored.

Black or African American↗

PubMed1993

H7-resistant protein kinase C substrates in two-dimensional gels of proestrous rat anterior pituitary gland.

The presence of Ca(2+)-dependent and -independent protein kinase C (PKC) activities which were stimulated by phorbol-12,13-dibutyrate (PDBu) and sensitive to the kinase inhibitor staurosporine (IC50 values approx. 100 nM) was demonstrated in proestrous rat anterior pituitary gland. These PDBu-induced activities were completely abolished by the PKC-specific inhibitors Ro31-8220 and GF109203X (3 microM). The Ca(2+)-independent activity was more resistant (IC50 = 61 microM) to the kinase inhibitor H7 than the Ca(2+)-dependent activity (IC50 approx. 20 microM), however, this (unusual) resistance to H7 was not observed in the brain regions, hypothalamus and hippocampus. Possible substrates for the Ca(2+)-independent PKC in anterior pituitary were identified by two-dimensional gel electrophoresis and autoradiography following incubation in vitro with [32P]phosphate and 300 nM PDBu +/- 300 nM staurosporine or 30 microM H7. The phosphorylation of six proteins (16, 16, 25, 36, 65 and 69 kDa) was found to be stimulated by PDBu and inhibited by staurosporine, but not H7, in whole tissue, and another two such phosphorylated proteins (each 76 kDa) were observed in microsomal subcellular fractions. These phosphoproteins may be substrates for an H7-resistant PKC isoform previously shown to mediate a number of cellular responses in rat anterior pituitary gland.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1993

Characterisation of protein kinase C isoforms and enzymic activity from the alpha T3-1 gonadotroph-derived cell line.

Western blots of alpha T3-1 cell extracts were immunostained with antibodies specific for various protein kinase C (PKC) isoforms. These revealed the presence of PKC types alpha, epsilon and zeta, but beta, gamma, delta and eta were not detected. The potency with which partially-purified cytosolic PKC from alpha T3-1 cells was activated by phorbol 12,13-dibutyrate (PDBu), mezerein and 1,2-dioctanoyl-sn-glycerol was assessed in the presence and absence of Ca2+. The inhibitors staurosporine, K252a, H7, GF109203X and Ro 31-8220 were tested on basal activity, PDBu-induced activity and Ca(2+) + PDBu-induced kinase activity. Each inhibitor showed distinct differences in their IC50 values under the three conditions, suggesting that these inhibitors may exhibit different potencies on the PKC isoforms present in alpha T3-1 cells. Although histone IIIs was used as the phosphate acceptor for most of these experiments, the efficiency of alpha, epsilon and zeta peptide and GS peptide substrates were also determined, with epsilon peptide giving the greatest activity in the presence of PDBu or Ca2+. Each substrate displayed a different pattern of activation under the conditions tested. Overall, the findings suggest that 3 or more PKC isoforms with varying specificities are present in gonadotroph-derived alpha T3-1 cells and that the contribution of each isoform should be considered when these cells are used in models of anterior pituitary cell function where PKC is involved.

Amino Acid Sequence↗

PubMed1993

A structural basis for sequence comparisons. An evaluation of scoring methodologies.

A residue-exchange matrix has been derived that is suitable for comparison of amino acid sequences. This matrix is based on the tabulation of 207,795 amino acid replacements observed in 65 homologous sets of structurally aligned three-dimensional structures (235 proteins). The majority of the data is from structural comparisons where there is between 15 and 40% sequence identity. As a result, a scoring matrix such as the one devised here should provide a sensitive basis for the comparison of amino acid sequences and the search for homologous sequences in amino acid databases. In order to assess the value of this matrix we have made a comparative analysis with 12 other published scoring matrices that have been used for the alignment of protein amino acid sequences. We find that the matrix derived here is among the better performers in terms of alignment significance, detection of homologous sequences and the accuracy of alignments.

Amino Acid Sequence↗

PubMed1993

Evidence for a distinct H7-resistant form of protein kinase C in rat anterior pituitary gland.

Inhibition of phorbol 12,13-dibutyrate-induced protein kinase C (PKC) activity from rat midbrain, anterior pituitary and a number of other tissues, as well as COS 7 cells, was studied in vitro. In anterior pituitary, Ca(2+)-independent activity was notably resistant to H7 but sensitive to staurosporine and Ro 31-8220. All Ca(2+)-dependent activity was sensitive to these three inhibitors. Mezerein and 1,2-dioctanoyl-sn-glycerol also activated this H7-insensitive PKC from anterior pituitary. The distribution of this activity, prominently expressed in pituitary and perhaps also lung, and its characteristic resistance to H7 but not other inhibitors, does not obviously correlate with that of any of the well-characterised PKCs, and may reflect either a novel or a modified isoform.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1993

Evidence that protein kinase C alpha has reduced affinity towards 1,2-dioctanoyl-sn-glycerol: the effects of lipid activators on phorbol ester binding and kinase activity.

The effect of 1,2-diacylglycerols on specific binding of [3H]phorbol 12,13-dibutyrate to cytosolic protein kinase C (PKC) was investigated in tissues reported to contain different proportions of PKC isoforms. In lung, frontal cerebral cortex and cerebellum cytosols (enriched in PKC alpha, beta and gamma, respectively) displacement of specific binding by phorbol 12,13-dibutyrate or diacylglycerols containing unsaturated acyl chains was of similar potency for each tissue. A range of 1,2-diacylglycerols containing saturated acyl chains exhibited varying affinities for [3H]phorbol 12,13-dibutyrate binding sites in each tissue; defining an optimal acyl chain length of around 14 carbons in each case. However, the affinities of saturated diglycerides were consistently lower in lung cytosol than in frontal cerebral cortex and cerebellum cytosols, with the greatest differences occurring at lower acyl chain lengths, especially with 1,2-dioctanoyl-sn-glycerol. Furthermore, a mixed micelle assay of PKC activity showed that 1,2-dioctanoyl-sn-glycerol displayed reduced potency at PKC alpha partially-purified from COS 7 cell cytosol compared to the mixture of PKC isoforms present in rat midbrain cytosol. Both low potency of 1,2-dioctanoyl-sn-glycerol as a displacer of [3H]phorbol 12,13 dibutyrate binding and the ability of arachidonic acid to act as an allosteric enhancer of binding, correlated with the proportional PKC alpha content of a range of tissues reported in the literature. In PKC enzyme activity assays, 1,2-dioctanoyl-sn-glycerol, but not phorbol 12,13-dibutyrate, was correspondingly a much poorer activator of PKC alpha from COS 7 cells than of the broad consensus of isoforms in rat midbrain. When alpha and beta isoforms were extensively-purified on DEAE-cellulose then hydroxyapatite, both the low affinity of 1,2-dioctanoyl-sn-glycerol for [3H]phorbol 12,13-dibutyrate binding sites and their allosteric regulation by arachidonic acid were confirmed to be characteristic of the alpha rather than the beta isoforms.

Amino Acid Sequence↗

PubMed1993

Alignment and searching for common protein folds using a data bank of structural templates.

We introduce an approach to protein comparisons in which tertiary-structure information is exploited in the alignment of a protein sequence of known tertiary structure, or an aligned set of sequences of known homologous structures, with one or more sequences. The local tertiary environments of residues in the one or more three-dimensional structures (defined in terms of residue accessibility to solvent, secondary structure and hydrogen bonding) are used to select position-specific amino acid substitution scores and produce a scoring template suitable for aligning sequences or searching sequence data banks. The amino acid substitution scores have been accumulated from 72 families of protein structures in which the observed substitutions have been classified according to features of the local structure. Hence, the value attributed to a particular amino acid interchange in the template is not a constant, but is dependent upon the environmental context in which that substitution has occurred. We have used these structural templates to align proteins, as well as to search an amino acid sequence data bank for proteins having a similar fold. Indeed, a database of templates that corresponds to both unique structures and aligned homologous structures from the Brookhaven Protein Data Bank has been produced. A new sequence can be searched against the database of templates in order to identify a similar tertiary fold even if the sequence is not significantly similar to any proteins of known three-dimensional structure.

Amino Acid Sequence↗

PubMed1993

Functional expression of 5-HT1c receptor cDNA in COS 7 cells and its influence on protein kinase C.

Two subtypes of receptors for serotonin (5-hydroxytryptamine; 5-HT) are known to stimulate inositol (1,4,5)-trisphosphate production, the 5-HT1c and 5-HT2 receptors. In this study we investigated the ability of 5-HT1c receptors, transiently expressed in COS 7 cells, to functionally interact with protein kinase C-alpha, the indigenous (phorbol ester-responsive) isoform of the enzyme in those cells. Serotonin caused translocation of the [3H]phorbol 12,13-dibutyrate (PDBu) binding site of PKC-alpha from the cytosolic to the membrane fraction in a Ca(2+)-dependent manner which was prevented by the 5-HT1c receptor antagonist mianserin. The lipid activators of PKC, PDBu and 1,2-dioctanoyl-sn-glycerol (DOG) also caused translocation, but through a mechanism apparently quite independent of Ca2+.

Animals↗

PubMed1993

Fragment ranking in modelling of protein structure. Conformationally constrained environmental amino acid substitution tables.

Conformationally constrained environment-dependent amino acid residue substitution tables have been constructed from a database comprising 33 homologous families of protein sequences aligned on the basis of their three-dimensional structures. Residues are allotted to one of 216 (or 54) classes of combinations of structural features. These include nine main-chain conformation classes, three classes of side-chain accessibility and eight (or two) classes of side-chain involvement in three types of hydrogen bond. Seven different main-chain conformational classes outside of regions of regular structure were identified in an analysis of the distributions of phi-psi torsion angles in 84 high-resolution crystallographic structures. Residue substitutions at equivalent positions in the structural alignments are included where the main-chain conformational class is conserved. Frequency data in the form of 216 (or 54) environment specific (20 x 20 residue type) matrices are then converted to probabilities. Two smoothing regimes incorporating entropy-driven weights were applied to the set of 54 tables. Predicted residue substitutions have been generated for individual residue positions in beta-hairpins and the hypervariable regions of the immunoglobulins. These have been compared with the observed sequence variation at the same positions using rank correlation methods. Measurements of chi 2 distances demonstrate the considerable improvement in predictive power at key residue positions identified from interactive graphics studies when compared to the Dayhoff MDM250 mutation matrix. An illustrative example is given of an application of the method in the ranking of loop fragments in model building studies of structurally variable regions in two subtilisins. A combined template scoring procedure is found to be 26-fold more discriminatory than the Dayhoff matrix. The success rate is approximately 85%.

Amino Acid Sequence↗