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Biomedical subjects

M I Khan

Publications and source records attributed to M I Khan.

At least 37 records · Page 2Linked to original sources
PubMed1999

Amplification of Proteus mirabilis chromosomal DNA using the polymerase chain reaction.

A Proteus mirabilis-specific polymerase chain reaction (PCR) was developed and standardized. The origin of the primers was a recombinant clone that contained P. mirabilis-specific Hind III fragment DNA of 3.5-kilobase pairs. Based on the sequence data of P. mirabilis recombinant clone, two primers designated MMKAP 1 and MMKAP 2 were synthesized for use in the PCR. A P. mirabilis-specific 3.5-kb pair DNA product was amplified by the primers from 18 strains of P. mirabilis, but not from other Protease species and bacteria. The minimum amount of target DNA detected by P. mirabilis PCR was 10 fg using ethidium bromide/ultraviolet exposure of gels or Southern blot hybridization with a P. mirabilis recombinant DNA probe.

Bacteria↗
PubMed1999

Production, purification and characterization of a constitutive intracellular alpha-galactosidase from the thermophilic fungus Humicola sp.

The thermophilic fungus Humicola sp constitutively produces intracellular alpha-galactosidase (1.33 U mg-1 protein) within 48 h at 45 degrees C in shaken flasks, when grown in a medium containing 7% wheat bran extract as a carbon source and 0.5% yeast extract as a nitrogen source. The enzyme has been purified to homogeneity by ultrafiltration, ethanol precipitation, DEAE cellulose and Sephacryl S-300 chromatography with a 124-fold increase in specific activity and 29.5% recovery. The molecular weight of the enzyme is 371.5 kDa by gel filtration on Sephacryl S-300 and 87.1 kDa by SDS-polyacrylamide gel electrophoresis. The enzyme has an optimum temperature of 65 degrees C and an optimum pH of 5.0. Humicola alpha-galactosidase is a glycoprotein with 8.3% carbohydrate content and is acidic in nature with a pI of 4.0. The KmS for p-nitrophenyl-alpha-D-galactopyranoside, O-nitrophenyl-alpha-D-galactopyranoside, raffinose and stachyose are 0.279, 0.40, 1.45 and 1.42 mM respectively. The enzyme activity was strongly inhibited by Ag+ and Hg2+. D-Galactose inhibited alpha-galactosidase competitively and the inhibition constant (Ki) for galactose was 11 mM.

Ascomycota↗
PubMed1999

Purification and characterization of a cell-surface lectin (Lectin II) from Agrobacterium radiobacter NCIM 2443.

A lectin was isolated from Agrobacterium radiobacter cell surface and purified. It is a monomer of 40 kDa as shown by SDS-PAGE. The lectin has a pI of 9.15 and amino acid composition of the lectin shows that 44% of the amino acids are hydrophobic. The lectin agglutinates rabbit erythrocytes but does not agglutinate human erythrocytes. It does not show specificity for monosaccharides except for D-glucosamine. Fetuin and its N-linked glycopeptide also inhibit the activity of the lectin but greater inhibition is shown by locust bean gum and Nicotiana tobaccum (tobacco) tissue extracts.

Amino Acids↗
PubMed1999

Crystallization and preliminary X-ray studies of the basic lectin from the seeds of Artocarpus hirsuta.

The basic lectin from Artocarpus hirsuta specific towards methyl alpha-galactose has been purified and crystallized using the hanging-drop vapour-diffusion method with ammonium sulfate as precipitant. Three different crystal forms, orthorhombic I, orthorhombic II and hexagonal, were grown under the same crystallization conditions. The orthorhombic forms belonged to space group P212121 with unit-cell dimensions a = 92.9, b = 99.8, c = 166. 2 A and a = 89.9, b = 121.9, c = 131.6 A, respectively. The unit-cell dimensions of the hexagonal form were a = b = 84.1 and c = 271.7 A and the space group was P6122.

Crystallization↗
PubMed1998

Characterization of a novel exo-N-acetyl-beta-D-glucosaminidase from the thermotolerant Bacillus sp. NCIM 5120.

An exo-N-acetyl-beta-d-glucosaminidase from the thermotolerant Bacillus sp. NCIM 5120 was purified to homogeneity by chromatography on CM-cellulose, Sephacryl S-300 and phenyl-Sepharose. The enzyme has a Mr of 230000 as determined by size exclusion chromatography on Sephacryl S-300/Sephadex G-200 and exhibited a relative subunit Mr of 60000 on denaturing gel electrophoresis. It is a neutral protein with a pI of 6.79. The optimum pH and temperature for the enzyme activity are 6.0 and 70 degreesC, respectively. Determination of the reaction stereochemistry indicates that the enzyme is a retaining glycosidase with the beta anomer of GlcNAc formed as the initial product. Determination of the energy of activation with different leaving groups (p-nitrophenol and 4-methyl-umbelliferone) reveals that the enzyme exhibits a biphasic Arrhenius plot with two characteristic energy of activation with an inflection temperature of 50 degreesC. The activation energy at temperatures below the inflection point was found to be higher than that above the inflection point. The energy of activation for 4-Me-Umb-beta-d-GlcNAc was higher at temperatures below the inflection point than for pNP-beta-d-GlcNAc (60.3 and 43.2 kJ mol-1, respectively). It hydrolyzes specifically, terminally linked beta(1-4) GlcNAc residues from the non-reducing end of oligosaccharides. Comparative studies on the hydrolysis of chito-oligosaccharides by the exo-N-acetyl-beta-d-glucosaminidase indicates that chitobiose is the best substrate with a Km and kcat of 0.34 mM and 24 microoff min-1mg-1, respectively. It also exhibits strict substrate specificity with respect to the glycone substitution as well as anomeric linkage.

Acetylglucosaminidase↗
PubMed1998

Thermodynamics of ligand (substrate/end product) binding to endoxylanase from Chainia sp. (NCL-82-5-1): isothermal calorimetry and fluorescence titration studies.

The binding of xylo-oligosaccharides to Chainia endoxylanase resulted in a decrease in fluorescence intensity of the enzyme with the formation of 1:1 complex. Equilibrium and thermodynamic parameters of ligand binding were determined by fluorescence titrations and titration calorimetry. The affinity of xylanase for the oligosaccharides increases in the order X2<X3<X4</=X5. Contributions from the enthalpy towards the free energy change decreased with increasing chain length from X2 to X4, whereas an increase in entropy was observed, the change in enthalpy and entropy of binding being compensatory. The entropically driven binding process suggested that hydrophobic interactions as well as hydrogen bonds play a predominant role in ligand binding.

Actinomycetales↗
PubMed1998

alpha-Galactoside binding lectin from Artocarpus hirsuta: characterization of the sugar specificity and binding site.

The hemagglutinin from the seeds of Artocarpus hirsuta was purified to homogeneity by ion-exchange chromatography on DEAE-cellulose and CM-sephadex. The native protein of molecular mass 60,000 (gel filtration) is made up of two pairs of unidentical subunits, alpha and beta with molecular masses of 15,800 and 14,130. The lectin is basic in nature (pI 8.5) and a glycoprotein with neutral sugar content of 6.25%. Rabbit as well as human erythrocytes (A, B and O) are agglutinated by the lectin. The lectin activity is neither affected by Ca2+, Mg2+ or Mn2+ nor by EDTA. Methyl alpha-D-galactopyranoside, pNP-alpha-D-galactopyranoside and pNP-alpha-D-N-acetylgalactosamine are the best inhibitors of the lectin. 4-Methylumbelliferyl-alpha-galactopyranoside fluorescence was quenched on binding to A. hirsuta lectin. The sugar has two binding sites per tetramer of the lectin with a Ka of 3.5x105 M-1 at 25 degrees C. Chemical modification studies indicate that the net positive charge associated with epsilon-NH2 of lysine residues and the phenyl ring of tyrosine are essential for the sugar binding activity of A. hirsuta lectin.

Animals↗
PubMed1998

Cell death in the brain of the HTx rat.

The structural changes in the hydrocephalic brain suggest that cell death must be a significant factor in the loss of cortical mantle thickness and cell numbers. In the developing hydrocephalic brain a similar conclusion might also be reached. We present data suggesting that in the developing hydrocephalic HTx rat brain, there are at least two processes that underlie the structural deficit in the cortex. The first is abnormal development of the cortex probably resulting from a combination of the genetic defect and blockage of CSF circulation in the fetus. The second is a direct consequence of raised intracranial pressure and is manifested in a lack of development of glial cells in the neonatal brain and in the loss of myelination and neurones as pressure rises after birth, particularly after skull plate fusion. We find little evidence of increased cell death by apoptosis, recording instead a decrease in apoptotic index. There is evidence that necrosis must have occurred.

Animals↗
PubMed1998

Fluorimetric studies on saccharide binding to the basic lectin from Artocarpus hirsuta.

The binding of Artocarpus hirsuta lectin to galactose and its derivatives was examined by fluorescence spectroscopy. The intrinsic fluorescence intensity of the lectin was enhanced by 55% upon binding to methyl alpha-galactose without any change in the emission maximum (333 nm). 4-Methyl umbellifery alpha-galactopyranoside showed 100% quenching of its fluorescence intensity upon binding to the lectin without any shift in the emission maximum (373 nm). The association constant for the binding of the above sugars to the lectin decreases with increasing temperature. Methyl group in the alpha anomeric position of galactose enhanced the binding while that in the beta position reduced the binding to the lectin. Solute quenching studies of the lectin using acrylamide, potassium iodide and cesium chloride indicated that the tryptophan residues were fully accessible to the neutral quencher, while only partly accessible to the ionic quenchers.

Disaccharides↗
PubMed1998

Effect of the novel angiotensin II type 1 receptor antagonist L-158,809 on acute infarct expansion and acute anterior myocardial infarction in the dog.

OBJECTIVES: To assess the effect of the angiotensin II type 1 receptor (AT1-R) antagonist L-158,809 on acute infarct expansion and left ventricular (LV) function during acute anterior myocardial infarction. METHODS: Dogs were randomized to receive intravenous L-158,809 (0.1 mg/kg bolus and 0.6 microgram/kg/min infusion) or vehicle beginning 1 h after permanent left anterior descending coronary artery ligation and continued for 48 h. In vivo LV remodelling and function (quantitative echocardiography) and hemodynamics over 48 h, and postmortem remodelling after 48 h were measured. RESULTS: L-158,809 produced 90% to 100% inhibition of the angiotensin II pressor response during the infusions. With respect to percentage changes over the 48 h in vivo, compared with vehicle controls, L-158,809 decreased mean arterial pressure (-20 +/- 4 versus -9 +/- 2%, P = 0.03) and left atrial pressure (-38 +/- 5 versus 25 +/- 6%, P < 0.0001) but did not change heart rate. These unloading effects were associated with a smaller percentage increase in infarct expansion index (-5 +/- 7% versus 27 +/- 2%, P = 0.001) and LV diastolic volume (11 +/- 11% versus 52 +/- 6%, P = 0.008), less shape deformation, fewer apical aneurysms (0 versus 100%, P = 0.0003), better global ejection fraction (49 +/- 2% versus 39 +/- 2%, P = 0.005), less ST segment elevation and fewer Q waves. Also compared with vehicle controls, with L-158,809 postmortem infarct size (19.8 +/- 2.4% versus 50.4 +/- 4.7% of risk region, P = 0.0002) and expansion index (2.06 +/- 0.09 versus 2.76 +/- 0.18, P = 0.006) were less and thinning ratio greater (0.92 +/- 0.02 versus 0.60 +/- 0.05, P = 0.0001). CONCLUSIONS: The novel AT1-R antagonist L-158,809 produces significant AT1-R blockade, reduces LV loading, and effectively limits acute infarct expansion and early LV remodelling during canine myocardial infarction.

Analysis of Variance↗
PubMed1997

Purification and characterization of an extracellular lectin (Lectin I) from Agrobacterium radiobacter NCIM 2443.

A lectin from culture filtrate of Agrobacterium radiobacter NCIM 2443 is purified to homogeneity by ion exchange chromatography on a DEAE cellulose column followed by hydrophobic chromatography on phenyl sepharose and hydroxyapatite column chromatography. The protein (Lectin I) is a monomer of relative molecular mass 37,000, as determined by denaturing gel electrophoresis as well as size exclusion chromatography. Lectin I is stable at pH 5.0 and its isoelectric point is pH 4.0. Amino acid analysis reveals that acidic amino acids and glycine are predominant amino acids and cysteine is absent in the lectin. Chemical modification of tryptophan residues causes more than 80% loss of haemagglutination activity of the lectin and 60% loss of activity is caused by modification of carboxyl groups. Lectin I agglutinates rabbit erythrocytes but does not agglutinate human A, B and O types of erythrocytes. It is specific for N-acetyl D-glucosamine, chitobiose, pNP-beta-mannoside as well as high mannose type glycopeptides. The relative inhibition by disaccharides, oligosaccharides and glycoproteins indicates that Lectin I recognizes Man3-GlcNAc-GlcNAc core carbohydrate structure of asparagine linked glycopeptides. Tobacco tissue extracts also inhibit the haemagglutination activity of Lectin I.

Animals↗
PubMed1997

Multiplex PCR for avian pathogenic mycoplasmas.

Mycoplasma infections are of great concern in avian medicine, because they cause economic losses in commercial poultry production. A multiplex polymerase chain reaction (PCR) was optimized to simultaneously detect four pathogenic species of avian mycoplasmas. Four sets of oligonucleotide primers specific for Mycoplasma gallisepticum (MG), M. synoviae (MS), M. meleagridis (MM) and M. iowae (MI) were used in the test. By using agarose gel electrophoreses for detection of the PCR-amplified DNA products, the sensitivity of detection was between 1 pg for MG, 1 pg for MS, 100 fg for MM and 100 pg for MI after 35 cycles of PCR. Similar sensitivity of these primers was achieved with broth cultures of these four organisms.

Animals↗
PubMed1997

Rapid estimation of spoilage bacterial load in aerobically stored meat by a quantitative polymerase chain reaction.

We report a quantitative PCR which utilizes primers from a conserved 23S rDNA sequence identified in nine different spoilage bacteria commonly present in meat. The PCR detected the spoilage bacteria by amplifying a specific 207 bp sequence from their chromosomal DNA. Quantification of PCR product by electrochemiluminescence revealed that the concentration of the amplified product was dependent on cycle number and the initial number of bacteria present in the sample. Statistical analysis of the results indicated a correlation coefficient of 0.94 (P < 0.001) between aerobic plate count and QPCR luminosity units.

Bacteria↗
PubMed1997

Effects of hydrocephalus on nitric oxide neurones in the brain of the HTx rat.

The effect of hydrocephalus on the nitric oxide system in the brain of the HTx rat brain was investigated. We used NADPH-diaphorase histochemistry to stain neurones containing the enzyme, nitric oxide synthase. Comparison of normal and hydrocephalic HTx rat brains showed a reduction in stained neurones and a greatly depressed staining intensity in those that were labelled in the hydrocephalic brain. Also, there was an apparent reduction in number and length of radiating fibres from stained neurones. In addition to these changes we also observed a clear association between stained neurones and the microvasculature of the brain. This suggests that changes in the activity of these neurones may have a significant impact on the normal perfusion of the brain.

Animals↗