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Biomedical subjects

M I Khan

Publications and source records attributed to M I Khan.

At least 91 records · Page 5Linked to original sources
PubMed1989

Differentiation of the vaccine F-strain from other strains of Mycoplasma gallisepticum by restriction endonuclease analysis.

The electrophoretic patterns of the nucleic acids (DNA) of Mycoplasma gallisepticum strains digested with the restriction enzymes Bam HI, Eco RI and Hind III were useful for differentiating the vaccine F-strain from other strains of M. gallisepticum. The procedure was more sensitive than the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) technique. The vaccine F-strain, represented by cultures designated F-K810 and F-F2F10 was clearly differentiated from other strains of M. gallisepticum. This procedure may be useful in field studies to determine if the vaccine strain will replace wild-type M. gallisepticum in commercial layers.

Bacterial Vaccines↗
PubMed1988

Interactions of concanavalin A with asparagine-linked glycopeptides: formation of homogeneous cross-linked lattices in mixed precipitation systems.

We have previously shown that certain oligomannose and bisected hybrid type glycopeptides are bivalent for binding to concanavalin A (Con A) [Bhattacharyya, L., Ceccarini, C., Lorenzoni, P., & Brewer, C. F. (1987) J. Biol. Chem. 262, 1288-1293]. Each glycopeptide gives a quantitative precipitation profile with the protein which consists of a single peak that corresponds to the binding stoichiometry of glycopeptide to protein monomer (1:2). We have shown that the affinities of the primary and secondary sites of the glycopeptides influence their extent of precipitation with the lectin [Bhattacharyya, L., & Brewer, C. F. (1988) Eur. J. Biochem. (in press)]. In the present study, we demonstrate that equimolar mixtures of any two of the glycopeptides result in a quantitative precipitation profile which shows two protein peaks. Using radiolabeled glycopeptides, the precipitation profiles of the individual glycopeptides were determined. The results show that each glycopeptide forms its own precipitation profile with the protein which is independent of the profile of the other glycopeptide. For mixtures containing an equimolar ratio of two glycopeptides, the glycopeptide with lower affinity shows a precipitation maximum at a lower concentration than the one with higher affinity. However, this can be reversed by increasing the ratio of the lower affinity glycopeptide in the mixture. Thus, the relative precipitation maxima of the glycopeptides are determined by mass-action equilibria involving competitive binding of the two carbohydrates to the protein. These equilibria, in turn, are sensitive to the relative amounts and affinities of the carbohydrates at both their primary and secondary sites.(ABSTRACT TRUNCATED AT 250 WORDS)

Asparagine↗
PubMed1988

Ubiquitin-metallothionein fusion protein expression in yeast. A genetic approach for analysis of ubiquitin functions.

We have established a Saccharomyces cerevisiae genetic system that expresses the fusion protein ubiquitin-metallothionein. We have evaluated the effects of amino-terminal ubiquitination of metallothionein on the stability and function of metallothionein. The fusion protein of wild type ubiquitin and metallothionein was rapidly processed in vivo to release free ubiquitin and metallothionein. Site-directed mutants of ubiquitin-metallothionein expressed in yeast were used to study the specificity of the (alpha-NH2-ubiquitin) protein endopeptidases. The data suggest that amino-terminal ubiquitination is not a signal for the proteolysis of yeast metallothionein in yeast. We have also discovered that expression of selected ubiquitin mutants blocked the growth of yeast. The data suggest that in addition to its function as a proteolytic signal, ubiquitination of proteins plays multiple roles in the cell.

Copper↗
PubMed1988

Binding of N-dansylgalactosamine to winged-bean tuber lectin: studies by fluorescence quenching titrations.

The winged-bean tuber lectin binds to N-dansyl(5-dimethylaminonaphthalene-1-sulphonic acid)galactosamine, leading to a 12.5-fold increase in dansyl fluorescence with a concomitant 25 nm blue-shift in the emission maximum. The enhancement of fluorescence intensity was completely reversed by the addition of methyl alpha-galactopyranoside. The lectin has two binding sites per molecule for this fluorescent sugar and an association constant of 2.59.10(5) M-1 at 25 degrees C. The binding of N-dansylgalactosamine to the lectin shows that it can accommodate a large hydrophobic substituent on the C-2 carbon of D-galactose. Studies with other sugars indicate that a hydrophobic substituent with alpha-conformation at the anomeric position increases the affinity of binding. The C-4 and C-6 hydroxyl groups are also critical for sugar binding to this lectin.

Binding Sites↗
PubMed1988

Antigenic analysis of three strains of Mycoplasma meleagridis of varying pathogenicity.

Three strains of Mycoplasma meleagridis of varying pathogenicity were characterized by conventional serologic methods and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The three strains (RY-39A, MC-25B, and RY-39C) did not differ by cross agglutination, immunofluorescence and immunodiffusion tests. However, the strains differed in their protein patterns as determined by SDS-PAGE.

Animals↗
PubMed1988

Nimodipine treatment in poor-grade aneurysm patients. Results of a multicenter double-blind placebo-controlled trial.

A multicenter, randomized placebo-controlled double-blind trial of nimodipine in poor-grade aneurysm patients was carried out in 17 Canadian hospitals. Of 188 patients enrolled in the trial, 32 were excluded for protocol violations and two were excluded due to statistical considerations, leaving 154 patients for valid outcome analysis. Nimodipine treatment was associated with a significantly better outcome (p less than 0.001): 21 (29.2%) of 72 nimodipine-treated patients had a good outcome at 3 months after subarachnoid hemorrhage (SAH) compared to eight (9.8%) of 82 placebo-treated patients. Delayed ischemic deficits from vasospasm alone were significantly less frequent in the nimodipine group (p less than 0.05) with permanent deficits occurring in five nimodipine-treated patients (6.9%) and in 22 placebo-treated patients (26.8%). Improvement in the good outcome rate and reduction in delayed ischemic deficits from vasospasm alone occurred in both Grade 3 and 4 patients, with no difference between nimodipine- and placebo-treated patients being found in Grade 5 patients. Repeat angiography after Day 4 was carried out in 124 patients. There was no significant difference in the incidence of moderate or severe diffuse spasm, which was seen in 64.3% of nimodipine-treated patients and 66.2% of placebo-treated patients. The authors conclude that nimodipine treatment in poor-grade patients with SAH results in an increase in the number of good outcomes and a reduction in the incidence of delayed neurological deterioration due to vasospasm. This effect occurs by a mechanism other than prevention of large-vessel spasm as visualized on angiography.

Adolescent↗
PubMed1987

Gene synthesis, expression, structures, and functional activities of site-specific mutants of ubiquitin.

To study the structure and function of ubiquitin we have chemically synthesized a ubiquitin gene that encodes the amino acid sequence of animal ubiquitin, inserting a series of restriction enzyme sites that divide the gene into eight "mutagenesis modules." A series of site-specific mutations were constructed to selectively perturb various regions of the molecule. The mutant genes were expressed in a large quantity of Escherichia coli, and the modified proteins were purified. To determine the structural effects of the amino acid substitutions, the solution structure of ubiquitin was investigated by two-dimensional NMR and each of the mutant proteins were screened for structural perturbations. With one exception, virtually no changes were seen other than at the point of mutation. Functional studies of the mutant proteins with the ubiquitin-activating enzyme E1 and in the reticulocyte protein degradation assay were used to identify regions of the molecule important to ubiquitin's activity in intracellular proteolysis.

Amino Acid Sequence↗
PubMed1987

Chemical synthesis and expression of a cassette adapted ubiquitin gene.

A gene encoding the yeast ubiquitin was chemically synthesized and expressed in yeast under regulatory control of the copper metallothionein (CUP1) promoter. The gene was assembled in a one-step ligation reaction from eight oligonucleotide fragments ranging in length from 50 to 64 nucleotides. To facilitate mutagenesis and gene fusion studies, eight unique 6-base-cutting restriction enzyme sites were placed in the reading frame which did not alter the encoded protein sequence or force the utilization of rare codons. In a copper-resistant yeast strain (CUP1r), expression of the gene was induced by copper to approximately 5% of the total yeast proteins, as determined by Coomassie-stained polyacrylamide gels. The protein, purified from yeast, reacted with ubiquitin-specific antibodies and was found to be biologically active in supporting ubiquitin-dependent protein degradation in vitro.

Amino Acid Sequence↗
PubMed1987

A vector for construction of gene libraries and the expression of heterologous genes in Saccharomyces cerevisiae.

We have constructed a convenient new vector, YEp-DE, for the construction of gene libraries and the expression of heterologous genes in Saccharomyces cerevisiae. The vector contains the yeast LEU2 gene, the 2 mu origin of replication, and a region from pUC18 that includes the ampr gene, the Escherichia coli origin of replication (ori), and the LacZ gene with multiple cloning sites. Five sites (Sac1, Sma1, BamH1, Sal1, Sph1) in this region are unique. This vector has advantages over similar yeast-E. coli shuttle vectors: small size (7291 bp, entirely sequenced), convenient cloning sites, and lacZ selection for detecting recombinant plasmids.

Cloning, Molecular↗
PubMed1986

Thermodynamic and kinetic studies on saccharide binding to soya-bean agglutinin.

The fluorescence of N-dansylgalactosamine [N-(5-dimethylaminonaphthalene-1-sulphonyl)galactosamine] was enhanced 11-fold with a 25 nm blue-shift in the emission maximum upon binding to soya-bean agglutinin (SBA). This change was used to determine the association constants and thermodynamic parameters for this interaction. The association constant of 1.51 X 10(6) M-1 at 20 degrees C indicated a very strong binding, which is mainly due to a relatively small entropy value, as revealed by the thermodynamic parameters: delta G = -34.7 kJ X mol-1, delta H = -37.9 kJ X mol-1 and delta S = -10.9 J X mol-1 X K-1. The specific binding of this sugar to SBA shows that the lectin can accommodate a large hydrophobic substituent on the C-2 of galactose. Binding of non-fluorescent ligands, studied by monitoring the fluorescence changes when they are added to a mixture of SBA and N-dansylgalactosamine, indicates that a hydrophobic substituent at the anomeric position increases the affinity of the interaction. The C-6 hydroxy group also stabilizes the binding considerably. Kinetics of binding of N-dansylgalactosamine to SBA studied by stopped-flow spectrofluorimetry are consistent with a single-step mechanism and yielded k+1 = 2.4 X 10(5) M-1 X s-1 and k-1 = 0.2 s-1 at 20 degrees C. The activation parameters indicate an enthalpicly controlled association process.

Acetylgalactosamine↗
PubMed1986

Thermodynamic and kinetic analysis of carbohydrate binding to the basic lectin from winged bean (Psophocarpus tetragonolobus).

A basic lectin (pI approximately 10.0) was purified to homogeneity from the seeds of winged bean (Psophocarpus tetragonolobus) by affinity chromatography on Sepharose 6-aminocaproyl-D-galactosamine. The lectin agglutinated trypsinized rabbit erythrocytes and had a relative molecular mass of 58,000 consisting of two subunits of Mr 29,000. The lectin binds to N-dansylgalactosamine, leading to a 15-fold increase in dansyl fluorescence with a concomitant 25-nm blue shift in the emission maximum. The lectin has two binding sites/dimer for this sugar and an association constant of 4.17 X 10(5) M-1 at 25 degrees C. The strong binding to N-dansylgalactosamine is due to a relatively positive entropic contribution as revealed by the thermodynamic parameters: delta H = -33.62 kJ mol-1 and delta S0 = -5.24 J mol-1 K-1. Binding of this sugar to the lectin shows that it can accommodate a large hydrophobic substituent on the C-2 carbon of D-galactose. Studies with other sugars indicate that a hydrophobic substituent in alpha-conformation at the anomeric position increases the affinity of binding. The C-4 and C-6 hydroxyl groups are critical for sugar binding to this lectin. Lectin difference absorption spectra in the presence of N-acetylgalactosamine indicate perturbation of tryptophan residues on sugar binding. The results of stopped flow kinetics with N-dansylgalactosamine and the lectin are consistent with a simple one-step mechanism for which k+1 = 1.33 X 10(4) M-1 s-1 and k-1 = 3.2 X 10(-2) s-1 at 25 degrees C. This k-1 is slower than any reported for a lectin-monosaccharide complex so far. The activation parameters indicate an enthalpically controlled association process.

Animals↗
PubMed1986

Fertility and pregnancy after operation for a prolactinoma.

Pituitary exploration was performed in 40 women with hyperprolactinaemic infertility and a prolactinoma was found in 37. After operation 26 (65%) of the 40 patients had a normal serum prolactin concentration; 31 women have so far had 49 pregnancies; in seven of these patients bromocriptine therapy was used after surgery. Eight of the remaining nine women were shown to be ovulating, but four of these subsequently decided not to attempt a pregnancy. The course and outcome of the 49 pregnancies were similar to those in normal women and no visual or endocrine complications occurred. Surgical removal of a prolactinoma often restores fertility, and may secure a safer pregnancy.

Adenoma↗