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C Robinson

Publications and source records attributed to C Robinson.

At least 37 records · Page 2Linked to original sources
PubMed2002

Organic material and the optical properties of the dark zone in caries lesions of enamel.

Sections of uncavitated natural caries lesions of human enamel were extracted with various solvents and examined by polarizing microscopy. After lipid extraction, the dark zone enlarged by 19-162% and its birefringence increased, while after protein extraction it shrank by 37-92% and became less birefringent. It is concluded that occlusion of submicroscopic pores by organic material largely accounts for the optical properties of the dark zone. The results are not consistent with occlusion of pores by reprecipitation of mineral. On the basis of previous work, organic material in this location could influence demineralization and remineralization.

Calcium↗
PubMed2002

Enamelin compartmentalization in developing porcine enamel.

The tissue compartmentalization of enamelin-processing products has been investigated in developing pig enamel using a sequential extraction procedure. Only trace amounts of enamelin-processing products were detected in simulated enamel fluid extracts, suggesting that enamelins are not solubilized in the matrix to any great extent. Subsequent phosphate buffer extraction desorbed and extracted several enamelin-processing products that were presumably bound to the mineral phase. A 35-kD processing product dominated the phosphate extract, suggesting that enamelin processing leads to an accumulation of this mineral-bound molecule. Dissociative extraction with urea subsequently extracted the remainder of the enamelin-processing products present. This material was presumably present in the tissue in an aggregated insoluble state. Several enamelin-processing products were only extracted by specific extraction procedures, suggesting that different enamelin-processing products are differentially compartmentalized. This may indicate that specific enamelin-processing products have different functions. In contrast to amelogenins, which are processed in the deeper tissue to generate products having a low affinity for the mineral, enamelin processing appears to produce products (those enamelins desorbed by phosphate buffer) that have a high affinity for the mineral. These products, appearing in the deeper enamel layers, may serve to influence crystal growth kinetics in the absence of any mineral-binding amelogenins.

Acetic Acid↗
PubMed2002

Inheritance pattern and elemental composition of enamel affected by hypomaturation amelogenesis imperfecta.

Hypomaturation amelogenesis imperfecta (AI) is characterized clinically by enamel of normal thickness that is hypomineralized, mottled, and detaches easily from the underlying dentin. Autosomal dominant, autosomal recessive, X-linked, and sporadic modes of inheritance have been documented. The present study investigated the elemental composition of the enamel of teeth from individuals demonstrating clinical hypomaturation AI from families representing three of these patterns of inheritance. The aim of the study was to determine if there was any commonality in microscopic phenotype of this defect between families demonstrating the various inheritance patterns. One section from each tooth was microradiographed and then viewed in a scanning electron microscope (SEM) equipped with an ultrathin window energy-dispersive x-ray spectroscopy (EDX) detector. In the SEM, prisms and constituent crystals in discrete areas appeared to be largely obscured by an amorphous material. EDX analysis showed enamel outside these areas to have a composition indistinguishable from control teeth. However, within these affected areas there was a large increase in carbon content (up to a fivefold increase). In some teeth there was also a detectable but smaller increase in the relative amounts of nitrogen or oxygen. The results suggest the defect in these teeth with a common clinical phenotype, irrespective of the pattern of inheritance, demonstrates a commonality in microscopic phenotype. The large increase in carbon content, not matched by an equivalent increase in nitrogen or oxygen, suggests a possible increased lipid content. In those teeth with elevated nitrogen levels there may also be retained protein.

Amelogenesis Imperfecta↗
PubMed2002

Characterization of a porcine amelogenin preparation, EMDOGAIN, a biological treatment for periodontal disease.

EMDOGAIN is derived from porcine developing enamel matrix and has been shown to facilitate regeneration of the periodontium, although its mechanism of action is unknown. The aim of the present study was to identify enamel matrix proteins and proteolytic enzymes present in EMDOGAIN and compare them with those extracted from developing porcine enamel itself. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), Western blotting, and zymography were used to identify the proteins present and to determine their enzyme activity. The results showed that developing enamel contained amelogenins, albumin, amelin, and enamelin. EMDOGAIN, however, contained only amelogenins. Both metalloendoproteases and serine protease activity were revealed in both EMDOGAIN and developing enamel. The roles of the amelogenin and enzyme components, if any, in periodontal regeneration are unknown.

Animals↗
PubMed2001

Distinct Albino3-dependent and -independent pathways for thylakoid membrane protein insertion.

The homologous proteins Oxa1, YidC, and Alb3 mediate the insertion of membrane proteins in mitochondria, bacteria, and chloroplast thylakoids, respectively. Depletion of YidC in Escherichia coli affects the integration of every membrane protein studied, and Alb3 has been shown previously to be required for the insertion of a signal recognition particle (SRP)-dependent protein, Lhcb1, in thylakoids. In this study we have analyzed the "global" role of Alb3 in the insertion of thylakoid membrane proteins. We show that insertion of two chlorophyll-binding proteins, Lhcb4.1 and Lhcb5, is almost totally blocked by preincubation of thylakoids with anti-Alb3 antibodies, indicating a requirement for Alb3 in the insertion pathway. Insertion of the related PsbS protein, on the other hand, is unaffected by Alb3 antibodies, and insertion of a group of SRP-independent, signal peptide-bearing proteins, PsbX, PsbW, and PsbY, is likewise completely unaffected. Proteinase K is furthermore able to completely degrade Alb3, but this treatment does not affect the insertion of these proteins. Among the thylakoid proteins studied here, Alb3 requirement correlates strictly with a requirement for stromal factors and nucleoside triphosphates. However, the majority of proteins tested do not require Alb3 or any other known form of translocation apparatus.

Arabidopsis Proteins↗
PubMed2001

Insertion of PsaK into the thylakoid membrane in a "Horseshoe" conformation occurs in the absence of signal recognition particle, nucleoside triphosphates, or functional albino3.

The photosystem I subunit PsaK spans the thylakoid membrane twice, with the N and C termini both located in the lumen. The insertion mechanism of a thylakoid membrane protein adopting this type of topology has not been studied before, and we have used in vitro assays to determine the requirements for PsaK insertion into thylakoids. PsaK inserts with high efficiency and we show that one transmembrane span (the C-terminal region) can insert independently of the other, indicating that a "hairpin"-type mechanism is not essential. Insertion of PsaK does not require stromal extract, indicating that signal recognition particle (SRP) is not involved. Removal of nucleoside triphosphates inhibits insertion only slightly, both in the presence and absence of stroma, suggesting a mild stimulatory effect of a factor in the translation system and again ruling out an involvement of SRP or its partner protein, FtsY. We, furthermore, find no evidence for the involvement of known membrane-bound translocation apparatus; proteolysis of thylakoids destroys the Sec and Tat translocons but does not block PsaK insertion, and antibodies against the Oxa1/YidC homolog, Alb3, block the SRP-dependent insertion of Lhcb1 but again have no effect on PsaK insertion. Because YidC is required for the efficient insertion of every membrane protein tested in Escherichia coli (whether SRP-dependent or -independent), PsaK is the first protein identified as being independent of YidC/Alb3-type factors in either thylakoids or bacteria. The data raise the possibility of a wholly spontaneous insertion pathway.

Amino Acid Sequence↗
PubMed2001

Functional characterization of recombinant chloroplast signal recognition particle.

The signal recognition particle (SRP) is a ubiquitous system for the targeting of membrane and secreted proteins. The chloroplast SRP (cpSRP) is unique among SRPs in that it possesses no RNA and is functional in post-translational as well as co-translational targeting. We have expressed and purified the two components of the Arabidopsis thaliana chloroplast signal recognition particle (cpSRP) involved in post-translational transport: cpSRP54 and the chloroplast-specific protein, cpSRP43. Recombinant cpSRP supports the efficient in vitro insertion of pea preLhcb1 into isolated thylakoid membranes. Recombinant cpSRP is a stable heterodimer with a molecular mass of approximately 100 kDa as determined by analytical ultracentrifugation, gel filtration analysis, and dynamic light scattering. The interactions of the components of the recombinant heterodimer and pea preLhcb1 were probed using an immobilized peptide library (pepscan) approach. These data confirm two previously reported interactions with the L18 region and the third transmembrane helix of Lhcb1 and suggest that the interface of the cpSRP43 and cpSRP54 proteins is involved in substrate binding. Additionally, cpSRP components are shown to recognize peptides from the cleavable, N-terminal chloroplast transit peptide of preLhcb1. The interaction of cpSRP43 with cpSRP54 was probed in a similar experiment with a peptide library representing cpSPR54. The C terminus of cpSRP54 is essential for the formation of the stable cpSRP complex and cpSPR43 interacts with distinct regions of the M domain of cpSRP54.

Amino Acid Sequence↗
PubMed2001

TatB and TatC form a functional and structural unit of the twin-arginine translocase from Escherichia coli.

In Escherichia coli, a subset of periplasmic proteins is exported via the twin-arginine translocation (Tat) pathway. In the present study, we have purified the Tat complex from E. coli, and we show that it contains only TatA, TatB, and TatC. Within the purified complex, TatB and TatC are present in a strict 1:1 ratio, suggesting a functional association. This has been confirmed by expression of a translational fusion between TatB and TatC. This Tat(BC) chimera supports efficient Tat-dependent export, indicating that TatB and TatC act as a unit in both structural and functional terms. The purified Tat complex contains varying levels of TatA, suggesting a gradual loss during isolation and a looser association. The molecular mass of the complex is approximately 600 kDa, demonstrating the presence of multiple copies of TatA, B, and C. Co-immunoprecipitation experiments show that TatC is required for the interaction of TatA with TatB, suggesting that TatA may interact with the complex via binding to TatC.

Base Sequence↗
PubMed2001

Private specificities can dominate the humoral response to self-antigens in patients with cryptogenic fibrosing alveolitis.

BACKGROUND: The pathogenetic mechanisms that underlie the interstitial lung disease cryptogenic fibrosing alveolitis (CFA) may involve an immunological reaction to unidentified antigens in the lung, resulting in tissue damage. METHOD: In order to identify the range of target autoantigens, we used expression cloning, employing serum from an index patient as the probe against an expressed cDNA library that was derived from a tumour cell line. We screened over 5 x 105 recombinants and obtained sequence information on three antigens that had provoked strong responses with immunoglobulin heavy chain class switching, presumably as a consequence of T-cell recognition. RESULTS: All of the antigens were identifiable by comparison with sequence data from the US National Center for Biotechnology Information. Alanyl tRNA synthetase (ATS) was picked on six occasions; five of these incidences reflected independent recombination events, indicating that the library was not biased. Antibodies to ATS (anti-PL-12) represent the most common reactivity that defines the antisynthetase syndrome, which is typically expressed as polymyositis, dermatomyositis and interstitial lung disease (ILD). The index patient never showed symptoms other than those associated with alveolitis, even though sera obtained from him over a period of 2 years contained antibodies with the same specificity. Autoantibodies to ATS were never detected in serial bleeds from 11 other patients with CFA, and neither did we detect antibodies to the other two antigens identified from the serum of the index patient. CONCLUSION: The humoral response in patients with CFA can be dominated by autoantibodies with private specificities. This suggests that the antibodies are epiphenomenal and are a secondary feature of tissue damage induced by some other mechanism.

Antibody Diversity↗
PubMed2001

Conformation of a purified "spontaneously" inserting thylakoid membrane protein precursor in aqueous solvent and detergent micelles.

Subunit W of photosystem II (PsbW) is a single-span thylakoid membrane protein that is synthesized with a cleavable hydrophobic signal peptide and integrated into the thylakoid membrane by an apparently spontaneous mechanism. In this study, we have analyzed the secondary structure of the pre-protein at early stages of the insertion pathway, using purified recombinant pre-PsbW. We show that the protein remains soluble in Tris buffer after removal of detergent. Under these conditions pre-PsbW contains no detectable alpha-helix, whereas substantial alpha-helical structure is present in SDS micelles. In aqueous buffer, the tryptophan fluorescence emission characteristics are intermediate between those of solvent-exposed and hydrophobic environments, suggesting the formation of a partially folded structure. If denaturants are excluded from the purification protocol, pre-PsbW purifies instead as a 180-kDa oligomer with substantial alpha-helical structure. Mature-size PsbW was prepared by removal of the presequence, and we show that this protein also contains alpha-helix in detergent but in lower quantities than the pre-protein. We therefore propose that pre-PsbW contains alpha-helical structure in both the mature protein and the signal peptide in nonpolar environments. We propose that pre-PsbW acquires its alpha-helical structure only during the later, membrane-bound stages of the insertion pathway, after which it forms a "helical hairpin"-type loop intermediate in the thylakoid membrane.

Chromatography, Gel↗
PubMed2001

Fluoride concentrations at and near the neonatal line in human deciduous tooth enamel obtained from a naturally fluoridated and a non-fluoridated area.

This study sought to obtain a precise profile of fluoride concentrations at and near the neonatal line in deciduous incisors and canines from the naturally fluoridated area (1.0--1.3 parts/10(6) F in drinking water) of West Hartlepool and the non-fluoridated area (less than 0.1 parts/10(6) F in drinking water) of Leeds in England. An abrasive microsampling method was used to determine the distribution of fluoride and phosphorus concentrations. The profile of fluoride concentrations in 100-microm layers before and after the neonatal line, that is, in the prenatal and postnatal enamel, were significantly higher in teeth from the fluoridated than non-fluoridated areas. It was concluded that the fact that the fluoride concentrations were about the same prenatally and postnatally in deciduous enamel obtained from the fluoridated and non-fluoridated areas indicates that fluoride enters the prenatal deciduous enamel and that it is transferred through the placenta.

Calcium↗
PubMed2001

Amelin extracellular processing and aggregation during rat incisor amelogenesis.

Amelin (also known as ameloblastin and sheathlin) is a recently described protein that is secreted by ameloblasts during enamel formation. Here, the extracellular distribution and processing of amelin during rat incisor amelogenesis were investigated by Western blot probing using anti-recombinant rat amelin antibodies. In addition, the solubility behaviour and aggregative properties of rat amelin were investigated using a sequential extraction procedure involving (1) extraction with simulated enamel fluid to extract proteins most likely to be soluble in vivo; (2) extraction with phosphate buffer to desorb proteins bound to enamel crystal surfaces; (3) extraction with sodium dodecyl sulphate (SDS) to extract proteins present as insoluble aggregates; followed by (4) a final acid demineralization step to release any remaining proteins. Proteins immunoreactive to the anti-amelin antibodies were detectable in secretory- and transition-stage enamel. Maturation-stage enamel appeared devoid of amelin. The largest immunoreactive protein detected migrated at 68 kDa on SDS gels, corresponding to the M(r) of nascent amelin. Other immunoreactive bands at 52, 40, 37, 19, 17, 16, 15, 14 and 13 kDa were presumably amelin processing products. The sequential extraction procedure revealed that the 68-, 52-, 40-, 37- and 13-kDa amelins were completely extracted under solution conditions similar to those reported to exist in vivo. In contrast, the 19-, 17- and 16-kDa amelins were only partially extracted, whilst the 15- and 14-kDa amelins could not be extracted with simulated enamel fluid. A proportion of the remaining 17- and 16-kDa amelins was desorbed from the enamel crystals with phosphate buffer and appeared to have been mineral-bound. The 15- and 14-kDa amelins and the remainder of the 17- and 16-kDa amelins were extracted with SDS only, suggesting that these species were present in vivo as an insoluble aggregate. The results provide additional information on amelin processing and degradation, and on how such processing influences the solubility and aggregative properties of amelin-derived proteins.

Ameloblasts↗
PubMed2001

Localised spontaneous regression in mesothelioma -- possible immunological mechanism.

Malignant mesothelioma (MM) is an aggressive tumor usually associated with asbestos exposure. Although it can remain stable for prolonged periods, it has not been described to spontaneously regress. MM tumors are thought to be immunogenic based both on animal studies and on the good responses in some humans treated with immunotherapy. Here we present a case of pleural MM in which a transient spontaneous regression was associated with tumor tissue infiltration with mononuclear cells and serological evidence of anti-MM reactivity. The patient's tumor eventually progressed and with this progression there was evidence of loss of serological reactivity to some, but not all, of her MM antigens. The patient survived for 20 months and, in contrast to her initial biopsy, no significant lymphoid infiltrate was detected in her MM tissue at post mortem examination.

Antibodies, Neoplasm↗
PubMed2001

A cost-benefit analysis of long-term management options for forests following contamination with 137Cs.

This paper provides a description of a cost-benefit analysis applied to determine the cost effectiveness, or otherwise, of nine management strategies potentially applicable to forests contaminated with 137Cs. The management strategies were considered singly and in a number of likely combinations. A management strategy was considered to be cost-effective if it resulted in a lower overall monetary detriment than was incurred if use of the contaminated forest was continued on a 'business as usual' basis. Only the banning of mushroom collection and restriction of public access proved to be cost-effective management strategies on the basis of this definition. However, even these strategies only proved cost-effective at high levels of 137Cs contamination, at which net savings in detriment in the form of public dose were achieved. Cost-effective savings of doses to forest workers were never achieved at any of the contamination levels considered in this study. It is suggested that novel alternative uses of contaminated forests are required which could provide added value to the standing crop in return for small increases in public and worker doses. One such use might be biofuel production.

Cesium Radioisotopes↗
PubMed2001

Investigation to evaluate and validate the Leeds in situ device for the study of enamel remineralisation in vivo.

OBJECTIVES: Recently, a novel device to generate dental plaque in situ on a removable human enamel surface was described. The device permitted the recovery of plaque intact and undisturbed on its enamel substrate. The aim of this investigation was to determine the utility and robustness of this model for analysis of the effects of therapeutics on both enamel remineralisation and on the overlying biofilm composition. METHODS: Enamel slices were taken from extracted sound human teeth, sterilised and a 'flat' area ground on each slice. An artificial lesion was formed within this area using acidified gel and the hardness of the enamel within the area of the lesion was determined at five sites using a 'Vickers' indenter. A nylon ring was then attached over the area of the lesion with cyanoacrylate and the excess enamel removed to form the completed device. Two devices were attached to the upper molars of 22 volunteers. Each volunteer was randomly assigned to receive either a fluoride containing (1500 ppm) or a fluoride free dentifrice. The devices were retained for a 4 week period whilst undertaking normal oral hygiene. All procedures were conducted according to GCP. After a 2 week break, the volunteers were fitted with two further devices, given dentifrice of the alternate type and the procedure repeated. Plaque from each device was harvested for microbiological analyses and the enamel subject to microhardness measurement. Then for each device the change in microhardness of the enamel within the lesion over the 4 week period was calculated. RESULTS: There were no significant differences in viable counts of total aerobic bacteria, mutans streptococci or lactobacilli, nor in acidic, aciduric or arginolytic populations in plaque from patients using the two different dentifrices. However, devices subjected to the fluoride containing dentifrice demonstrated a significantly greater increase in microhardness of the enamel (P<0.025). CONCLUSIONS: These results suggest that the novel in situ device is capable of measuring the effect of 1500 ppm fluoride on remineralisation of carious enamel over a 4 week period and is also well suited to determining concomitant effects on plaque ecology.

Adult↗