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J R Baker

Publications and source records attributed to J R Baker.

At least 127 records · Page 7Linked to original sources
PubMed1994

Characterization of the group B streptococcal hyaluronate lyase.

Hyaluronate lyase is one of several proteins secreted by group B streptococci which are believed to contribute to strain virulence. Characterization of the purified enzyme revealed that it degrades hyaluronan by a mechanism different from that of other previously studied hyaluronidases. Instead of randomly cleaving hyaluronan chains leading to a continuous decrease in average chain size, the group B streptococcal enzyme initially yields primarily unsaturated disaccharides. The observation that most of the free reducing ends generated during group B streptococcal hyaluronate lyase digestion are present in the unsaturated disaccharide units supports the conclusion that they are released primarily from the ends of the hyaluronan chains. Furthermore, the experimental evidence is consistent with a mode of action by which the enzyme initially makes a random cut in a hyaluronan chain and then processively moves along the chain releasing disaccharide units. Group B streptococcal hyaluronate lyase also slowly degrades chondroitin sulfate, and its desulfation greatly increases the reaction rate. A preferential cleavage of unsulfated residues is consistent with the observed extensive release of free chondroitin sulfate chains following very limited digestion of aggrecan from bovine nasal cartilage.

Animals↗
PubMed1994

The origins of parasitism in the protists.

The origins of parasitism among the protists are, like the group itself, polyphyletic. Probably the majority of present-day parasitic forms evolved from free-living ancestors which were ingested as part of the food of their hosts, though origins from ectoparasitic forms and via a phase of facultative parasitism are possibilities, particularly among the ciliated protozoa and (for ectoparasitism) the Kinetoplasta. Sporozoan parasites most probably developed via a stage which was ingested and became adapted to life in the host's gut. Further developments in parasitism involved deeper penetration into the host's tissues and the adoption of more than one host in the life cycle, thus avoiding entirely the potentially hazardous phase of existence outside the host.

Amoebida↗
PubMed1994

An analysis of the structure and antigenicity of different forms of human thyroid peroxidase.

Thyroid peroxidase (TPO) is an important enzyme in the production of thyroid hormone and one of the major autoantigens in autoimmune thyroid disease. The gene for human thyroid peroxidase encodes a single 933 amino acid polypeptide chain. However, several reports have suggested that it exists in both high- and low-molecular-weight forms and the exact structure of the native enzyme is not known. We examined the structure of TPO using two monoclonal antibodies against different portions of TPO, a polyclonal mouse antiserum raised against a 300 amino acid fragment of TPO and autoantibodies directed against TPO obtained from patients with autoimmune thyroid disease. Western blots performed under nonreducing conditions identified three bands of approximately 220-230 kDa and two bands of 105 and 110 kDa that appeared to be immunologic TPO. After reduction, the TPO activity migrated as a smear of bands from 105 to 110 kDa, suggesting that the higher molecular weight form of the enzyme is a disulfide-linked dimer. Patients with autoimmune thyroid disease showed higher rates of recognition of the dimer than the reduced monomer when serologic reactivity was analyzed by Western blots. Eighty-three percent (40 of 48) of patients with Graves' disease and 76% (34 of 45) of Hashimoto's disease patients recognized the dimer form of TPO, while 48% (23 of 48) of Graves' and 60% (27 of 45) of Hashimoto's patients recognized reduced monomer TPO, even though both forms were denatured with SDS. Antibodies against different portions of the TPO chain all bound to the 105 kDa bands, indicating that the TPO chain is not bisected during posttranslational processing.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1994

Characterization of human airway epithelial cell leukotriene A4 hydrolase.

We have previously shown that human airway epithelial cells contain leukotriene A4 (LTA4) hydrolase activity. To characterize this activity further, airway epithelial cells, cultured to confluence, were disrupted by sonication and were fractionated at 15,000 and 100,000 x g. Enzymatic activity was assessed by incubating fractions with 15 microM LTA4 at 37 degrees C for 15 min. LTA4 hydrolase activity was present in the 15,000 x g and the 100,000 x g supernatants and was inactivated by heating at 56 degrees C or by pronase, as is the case for neutrophil LTA4 hydrolase. However, the epithelial cell enzyme had a slower time course for product generation and demonstrated a different dose-response relationship to substrate when compared with the neutrophil. Kinetic analysis revealed nonlinear plots for epithelial data, most consistent with an enzyme that has multiple active sites. Immunoblotting, performed with anti-neutrophil LTA4 hydrolase antibody, recognized two bands in epithelial cell 15,000 x g supernatant (M(r) of 69,000 and 110,000-115,000). When resolved by gel filtration chromatography, only the M(r) 69,000 protein had enzymatic activity. Anion exchange chromatography of epithelial cell samples revealed that LTA4 hydrolase and aminopeptidase activity did not co-elute, whereas one of three peaks of aminopeptidase activity did co-elute in chromatograms of neutrophil samples. Immunoblots of proteolytic digests of partially purified M(r) 69,000 protein from epithelial cells and neutrophils revealed different immunoreactive bands. The digest of the M(r) 110,000-115,000 protein revealed no immunoreactive bands. Repeat kinetic analysis on 179-fold purified epithelial LTA4 hydrolase again revealed that it lacked significant aminopeptidase activity and retained its unique kinetic properties.(ABSTRACT TRUNCATED AT 250 WORDS)

CD13 Antigens↗
PubMed1994

Leukotriene A4 hydrolase in human bronchoalveolar lavage fluid.

We examined cell-free human bronchoalveolar lavage fluid (BALF) for enzymes of the 5-lipoxygenase pathway. BALF was obtained from six patients who were active smokers and six nonsmokers. Enzymatic activity in cell-free BALF was assessed by specific assays for leukotriene (LT) A4 hydrolase, 5-lipoxygenase, and LTC4 synthase using HPLC. Only LTA4 hydrolase enzymatic activity was found. This activity ranged from 101 to 667 when expressed as picomoles of LTB4 produced per milliliter BALF. Enzymatic activity in smokers vs nonsmokers was 484 +/- 120 vs 129 +/- 32 pmol LTB4/ml BALF (mean +/- SD, P < 0.0001). There were no leukotrienes found in BALF before assay. Immunoblot analysis revealed an immunoreactive band at a relative molecular mass of 69,000 D in all samples, consistent with LTA4 hydrolase, but no evidence of 5-lipoxygenase. BALF had greater LTA4 hydrolase activity per milligram of protein than neutrophil cytosol, epithelial cell cytosol, plasma, or serum. The synthesis of LTB4 was significantly increased when neutrophils were stimulated in BALF. These data indicate the selective presence of LTA4 hydrolase in BALF which is significantly increased in smokers. This enzyme in BALF may contribute to the inflammatory response in tobacco-related lung disease.

Adult↗
PubMed1994

Chemistry of the fructosamine assay: D-glucosone is the product of oxidation of Amadori compounds.

The chemistry of the fructosamine assay was studied by using the Amadori compound, N alpha-formyl-N epsilon-fructose-lysine (fFL), an analog of glycated lysine residues in protein. Previously (Clin Chem 1993;39:2460-5), we reported that free lysine was formed from fFL at 70% yield during incubation with alkaline nitroblue tetrazolium (NBT) under the conditions routinely used for the fructosamine assay (sodium carbonate buffer, pH 10.35 at 37 degrees C). Here, we show that D-glucosone is the primary carbohydrate oxidation product formed from Amadori compounds in the fructosamine assay. Glucosone, which decomposes under alkaline assay conditions with a half-life of < 30 min, reaches a maximum concentration of approximately 50% of the initial fFL concentration after 10 min of incubation. Like fFL, glucosone reduces NBT to the purple monoformazan dye, but its decomposition is not accelerated by the presence of NBT. The dicarbonyl-trapping reagent, aminoguanidine, inhibits the fructosamine assay by approximately 25% when fFL is the substrate, but by nearly 100% with glucosone as substrate. Studies with serum samples from diabetics and nondiabetics indicate that glucosone formation does not have a significant effect on the clinical usefulness of the fructosamine assay; however, corrections for glucosone formation may be required when the assay is used for estimating the extent of glycation of proteins.

Borohydrides↗
PubMed1993

Isolation and characterization of the chains of type V/type XI collagen present in bovine vitreous.

Previous studies show that the collagen fibrils of the mammalian vitreous humor are assembled largely from type II collagen with smaller amounts of type IX collagen and either type V or type XI collagen. In this paper, we report the separation of two chains of type V/type XI collagen from type II collagen by heparin-Sepharose chromatography. These chains were characterized by sequencing of selected cyanogen bromide or tryptic peptides with subsequent comparison of these sequences with cDNA-derived amino acid sequences of the alpha 1(V), alpha 1(XI), alpha 2(V), and alpha 2(XI) chains. The results show that vitreous fibrils are assembled from molecules containing the alpha 1(XI) and alpha 2(V) chains. These results, together with recent results from other laboratories, indicate that type V and type XI collagens are not separate collagen types but are part of a larger collagen family in which chains of both type V and type XI collagens participate in the formation of a variety of native molecules.

Amino Acid Sequence↗
PubMed1993

TSH beta subunit gene expression in human lymphocytes.

Production of neuroendocrine peptides by human lymphocytes is thought to facilitate control of the immune response. The presence of neuroendocrine peptide gene expression, specifically the TSH beta subunit gene, was studied in human lymphocytes using Northern blot analysis and polymerase chain reaction (PCR) techniques. Northern blot analysis of human lymphocyte RNA probed with a TSH beta cDNA probe failed to demonstrate TSH beta subunit steady state message levels. PCR amplification of lymphocyte-derived cDNA using TSH beta subunit complementary primers resulted in amplification of a .38 Kb DNA fragment, confirming expression and initial exonic splicing of TSH beta subunit gene exons 2 and 3 in human lymphocytes. Sequence analysis of this .38 Kb DNA fragment demonstrated conservation of exon borders after splicing (exons 2 and 3) and predicted an amino acid translation product similar, if not identical, to human TSH beta peptide sequence. Hybridization with a TSH beta subunit cDNA probe of PCR-amplified reverse-transcribed lymphocyte RNA suggested that: (1) the abundance of TSH beta subunit gene transcripts in human lymphocytes is less than the relative abundance in T3-treated pituitary; and (2) this messenger RNA may be modulated by the presence of certain thyromimetic compounds (T2, T3, TRIAC).

Amino Acid Sequence↗
PubMed1993

Rhythmicity associated with a high degree of short-term synchrony of motor unit discharge in man.

A rhythmic modulation of motor unit discharge at 17-24 Hz has been observed in the tibialis anterior muscle of subjects who display a high degree of short-term synchrony. Motor unit discharges were recorded both from extensor digitorum communis in the arm and tibialis anterior in the leg during voluntary isometric contractions. Time domain analysis of motor unit discharges in both muscles revealed a prominent central peak in the cross-correlograms that was interpreted as short-term synchrony. In tibialis anterior, but not in extensor digitorum communis, the primary, central peak was accompanied by secondary peaks. These were symmetrically or unilaterally present at an average time lag of 52 ms (range 37-68 ms) with respect to the primary peak. In both muscles other peaks occurred at time lags equivalent to the mean interspike interval of motor unit discharge (range 80-150 ms). In all subjects frequency domain analysis of the discharges of pairs of motor units revealed prominent peaks of coherence in the range 17-24 Hz. The actual frequency matched closely the reciprocal of the time lag interval between primary and secondary peaks of the cross-correlogram of the same motor units. Coherence was less evident for motor unit discharges in extensor digitorum communis and no prominent peaks occurred in the range 17-24 Hz. We propose that the secondary peaks and the rhythmicity may involve recurrent inhibition activated by the synchronous discharge of motor units. The presence of this pattern of discharge in tibilias anterior but not extensor digitorum communis would fit with recent animal and human work suggesting that recurrent inhibition is poorly represented in certain distal muscles of the forelimb.

Adult↗
PubMed1993

Mechanisms of peroxynitrite-induced injury to pulmonary surfactants.

Activated alveolar macrophages secrete both nitric oxide and superoxide in the alveolar lining fluid which combine rapidly to form peroxynitrite, a potent oxidizing agent capable of damaging lipids and proteins in biological membranes. Peroxynitrite (1 mM) plus 100 microM Fe3+EDTA inhibited calf lung surfactant extract (CLSE) from reaching a minimum surface tension below 10 mN/m on dynamic compression. Peroxynitrite and its by-products reacted with the unsaturated lipid components of CLSE, as evidenced by the appearance of conjugated dienes and thiobarbituric acid products, and damaged all surfactant proteins. A mixture of the hydrophobic proteins [surfactant protein B (SP-B) and surfactant protein C (SP-C)] exposed to peroxynitrite became incapable of lowering phospholipid minimum surface tension on dynamic compression. Exposure of SP-A to peroxynitrite decreased its ability to cause lipid aggregation and to act synergistically with SP-B and SP-C in lowering surface tension of surfactant lipids. Western blot analysis of SP-A exposed to peroxynitrite was consistent with fragmentation and polymerization of the 28- to 36-kDa triplet band, and amino acid analysis revealed the presence of significant levels of 3-nitro-L-tyrosine. We conclude that peroxynitrite and its reactive intermediates inhibit pulmonary surfactant function by lipid peroxidation and damaging surfactant proteins.

Animals↗
PubMed1993

Tissue specificity and serologic reactivity of an autoantigen associated with autoimmune thyroid disease.

A recent report has identified a new autoantigen called D1 that appears to be associated with Graves' ophthalmopathy and is expressed in the thyroid and eye muscle. To better characterize the tissue specificity and disease relevance of this antigen, we evaluated the expression of D1 RNA in various human tissues using a reverse transcriptase polymerase chain reaction assay. These studies indicate a wide tissue distribution of the messenger RNA for this antigen, including the thyroid, eye muscle, parathyroid, spleen, skeletal muscle, and uterus. There were variations in the relative amounts of specific message for D1 in the different tissues, with the uterus, thyroid, and eye muscle having the greatest amount of product per microgram of total RNA. A maltose binding protein-D1 fusion protein was expressed in Escherichia coli, purified, and used to assess serologic reactivity to D1 by Western blot. Autoantibodies to this antigen were noted in 19 of 24 (78%) of Hashimoto's disease patients, 26 of 41 (63%) of Graves' disease patients, and in 9 of 17 (53%) of normal controls. Sixty percent of Graves' disease patients with clinical ophthalmopathy had antibodies to D1, as did 63% of Graves' patients without signs or symptoms of clinical ophthalmopathy. There was no correlation between reactivity to D1 and either clinical measures of hyperthyroidism or antibody titers to thyroid peroxidase or thyroglobulin. The presence of autoantibodies to this antigen in patients with Hashimoto's disease, in Graves' disease patients without ophthalmopathy and in normal controls indicate that serologic recognition of this antigen is not restricted to patients with ophthalmopathy. In addition, the expression of messenger RNA for this antigen in multiple types of cells questions the tissue specificity of this autoantigen.

Adult↗
PubMed1993

Differential autoantibody responses to thyroid peroxidase in patients with Graves' disease and Hashimoto's thyroiditis.

In order to examine the specificity of the autoantibody response to thyroid peroxidase (TPO, previously identified as thyroid microsomal antigen) in autoimmune thyroid disease, we examined reactivity of sera from 45 Hashimoto's and 48 Graves' patients to native thyroid microsomes, denatured and reduced human TPO and several recombinant fragments of human TPO corresponding to amino acids 457-933 of the native protein. Both Graves' and Hashimoto's sera bound native, denatured and reduced TPO at significantly greater rate than normal controls, and no differences were noted between the two disorders in binding to these forms of the autoantigen. Binding was also noted to two recombinant fragments of TPO, corresponding to amino acids 513-633 and 633-933 in TPO. The frequency of autoantibodies to the TPO AA(633-933) region was not significantly different in Hashimoto's vs. Graves' disease patients (58% vs. 65% respectively), and appeared to relate to evidence of glandular inflammation in the Graves' patients (presence of anti-thyroglobulin antibodies and elevated anti-microsomal antibody levels). In contrast, antibodies to the TPO AA(513-633) fragment were significantly more common and of higher titer in Hashimoto's vs. Graves' disease patients, and did not correlate with any measure of glandular inflammation. These results identify two specific regions of TPO autoantibody binding and indicate that there are differences in the autoantibody response to TPO in Hashimoto's and Graves' diseases.

Adolescent↗
PubMed1993

Microalbuminuria in a middle-aged workforce. Effect of hyperglycemia and ethnicity.

OBJECTIVE: To determine the prevalence of microalbuminuria in a mixed, ethnic population and to find the extent that ethnic variation in microalbuminuria can be explained by abnormal glucose metabolism, obesity, hypertension, hypertriglyceridemia, and life-style factors. RESEARCH DESIGN AND METHODS: Urinary albumin concentrations were measured in 5467 middle-aged Maori, Pacific Islander, and European workers who participated in a health-screening survey of 46 New Zealand companies. Participants provided a first-voided, morning urine sample; had a 75-g oral glucose tolerance test; had weight, height, and blood pressure measured; and completed a self-administered questionnaire about past medical history and sociodemographic status. RESULTS: A significantly higher prevalence of microalbuminuria was found in individuals with new cases of diabetes mellitus (24.1%), in cases of diabetes mellitus previously diagnosed (20.6%), and in those with impaired glucose tolerance (16.1%) compared with nondiabetic individuals (4.0%). Moreover, in the general population, a piecewise linear relationship was detected between albuminuria and plasma glucose with significant changes of slope corresponding with 2 h plasma glucose concentrations (95% confidence interval) of 6.7 (6.4-7.0) and 9.2 (8.6-9.8) mM, respectively. After adjusting for sex, obesity, hypertension, hypertriglyceridemia, cigarette smoking, and heavy alcohol consumption in a multivariate model, glycemia was the most significant determinant of urinary albumin concentrations in all three ethnic groups. However, blood glucose concentrations did not completely explain the higher relative risk (95% confidence interval) of microalbuminuria in Maori (5.97; 4.48-7.78) and Pacific Islander (5.33; 4.13-6.87) workers compared with European workers. CONCLUSIONS: Of the variables investigated, hyperglycemia was the most important factor explaining the high prevalence of microalbuminuria in Maori and Pacific Islander workers compared with the European workers. However, only 14.9% of the variation in urinary albumin concentrations was found in our multivariate model, and we have speculated on the contribution of other factors such as diet and coexisting renal diseases.

Adult↗
PubMed1993

Asymptomatic bacteriuria in a multiracial workforce.

Prevalence of asymptomatic bacteriuria was determined in the first morning urine specimens from 5669 people who participated in a health screening survey of a local workforce. Higher age-standardized prevalences were observed in Maori women (18.0%), compared with Pacific Island women (9.8%) and European women (8.7%). Similarly, higher prevalences were observed in Maori men (3.2%), compared with Pacific Island men (0.7%) and European men (1.3%). After controlling for age and gender, the relative risk for asymptomatic bacteriuria in Maori people was estimated to be 2.22 times that for non-Maori people (associated 95% CI: 1.54-3.18). Significant associations of bacteriuria included a past history of myocardial infarction, a past history of kidney disease, absence of tertiary education, and impaired glucose tolerance. The most common pathogens, isolated from a subsample of 1660 participants, were Escherichia coli, 27 cases (50.9%), and Streptococcal sp, 14 cases (26.4%).

Adult↗
PubMed1993

Mechanism of fructosamine assay: evidence against role of superoxide as intermediate in nitroblue tetrazolium reduction.

We studied the chemistry of the fructosamine assay for glycated serum proteins by using the model Amadori compound N alpha-formyl-N epsilon-fructoselysine (fFL), an analog of glycated lysine residues in protein. Free lysine was formed at approximately 70% yield during a standard 20-min incubation of fFL with alkaline nitroblue tetrazolium (NBT) at 37 degrees C. Although superoxide dismutase (SOD; EC 1.15.1.1) and catalase (EC 1.11.1.6) decreased the yield of the product, monoformazan dye (MF+), the yield of MF+ was slightly greater under anaerobic than aerobic conditions, excluding a role for superoxide as an intermediate in the reduction of NBT during the fructosamine assay. SOD added to diabetic patients' sera at physiological concentrations also caused a significant (approximately 50%) inhibition of MF+ formation. This inhibition was reduced by addition of nonionic detergents, which contain organic peroxide inhibitors of SOD, to the fructosamine reagent. Overall, these data indicate that the Amadori compound is the direct reductant of NBT in the fructosamine assay and that superoxide is not an intermediate in the reaction. The inhibitory effects of SOD and catalase are most likely the result of oxygen regeneration in the assay mixture.

Catalase↗
PubMed1993

Albuminuria in people at least 40 years old: effect of alcohol consumption, regular exercise, and cigarette smoking.

We investigated the relation between albuminuria and life-style factors in 5670 people, ages 40 years and over, who participated in a health screening survey of a local workforce. The degree of albuminuria showed piecewise log-linear relationships with alcohol consumption and cigarette smoking, with changes in slope (and 95% confidence interval) corresponding with 5 (2, 8) g of alcohol/day and 10 (6, 14) cigarettes/day. After adjusting for age, gender, ethnicity, and other life-style variables, relative risks (95% confidence interval) of slight albuminuria for people consuming > 32 g of alcohol/day compared with nondrinkers, and for cigarette smokers compared with nonsmokers, were 1.74 (1.02, 2.98) and 1.37 (1.01, 1.88), respectively. However, there was no significant effect of exercise. We conclude that slight albuminuria is significantly associated with cigarette smoking and heavy alcohol consumption, consistent with its role as an index of risk of cardiovascular disease.

Adult↗