PubMed Health⌕ Search

Biomedical subjects

N Sharma

Publications and source records attributed to N Sharma.

At least 37 records · Page 2Linked to original sources
PubMed2001

Characterization of a galactose specific adhesin of enteroaggregative Escherichia coli.

A fimbrial adhesin was identified from an enteroaggregative Escherichia coli strain. The adhesin was purified to 740-fold by sequential chromatography on an affinity matrix and gel filtration column in the FPLC system. The homogeneity of the purified protein was established by analytical isoelectrofocussing (pI 7.25). The native adhesin appeared as a high-molecular-weight aggregative protein as revealed by gel filtration chromatography on Superose 12HR10/30 column. However, in sodium dodecyl sulfate-polyacrylamide gel electrophoresis the molecular weight of the adhesin was found to be 18 kDa and this was further confirmed by gel filtration chromatography on Superose 6HR 10/30 column presence of 6 M guanidine hydrochloride. The N-terminal 15-amino-acid sequence of the adhesin did not show homology with any of the previously reported fimbrial adhesins. The purified adhesin showed adhesion to human erythrocytes in the presence of Ca(2+) (5 mM). The optimum temperature and pH for the hemadhesion activity was found to be 25 degrees C and 6.5, respectively. The inhibition study clearly suggested that the binding site of the adhesin could recognize galactose as the specific sugar. The fluorescence of 4-methylumbelliferyl-alpha-D-galactopyranoside was quenched on binding to the adhesin and maximum reversal of fluorescence quenching was observed by competitive substitution titration with raffinose. The adhesin was found to contain one binding site per monomer for its specific sugar residue. The association constant and the free energy of binding were obtained as 3.98 x 10(5) M(-1) and -31.97 kJ/mol, respectively. The adherence of the bacteria to HEp-2 monolayer was inhibited in presence of galactose and this was further supported by a significant reduction in the bacterial adherence to the HEp-2 cells, pretreated with beta-D-galactosidase.

Adhesins, Escherichia coli↗
PubMed2001

Rpb4, a non-essential subunit of core RNA polymerase II of Saccharomyces cerevisiae is important for activated transcription of a subset of genes.

A major role in the regulation of eukaryotic protein-coding genes is played by the gene-specific transcriptional regulators, which recruit the RNA polymerase II holoenzyme to the specific promoter. Several components of the mediator complex within the holoenzyme also have been shown to affect activation of different subsets of genes. Only recently has it been suggested that besides the largest subunit of RNA polymerase II, smaller subunits like Rpb3 and Rpb5 may have regulatory roles in expression of specific sets of genes. We report here, the role of Rpb4, a non-essential subunit of core RNA polymerase II, in activation of a subset of genes in Saccharomyces cerevisiae. We have shown below that whereas constitutive transcription is largely unaffected, activation from various promoters tested is severely compromised in the absence of RPB4. This activation defect can be rescued by the overexpression of cognate activators. We have localized the region of Rpb4 involved in activation to the C-terminal 24 amino acids. We have also shown here that transcriptional activation by artificial recruitment of the TATA-binding protein (TBP) to the promoter is also defective in the absence of RPB4. Surprisingly, the overexpression of RPB7 (the interacting partner of Rpb4) does not rescue the activation defect of all the promoters tested, although it rescues the activation defect of the heat shock element-containing promoter and the temperature sensitivity associated with RPB4 deletion. Overall, our results indicate that Rpb4 and Rpb7 play independent roles in transcriptional regulation of genes.

Amino Acid Sequence↗
PubMed2001

Membrane-associated respiratory syncytial virus F protein expressed from a human rhinovirus type 14 vector is immunogenic.

Human rhinovirus (HRV) replicons have the potential to serve as respiratory vaccine vectors for mucosal immunization in humans. However, since many vaccine immunogens of interest are glycosylated, an important concern is whether HRV replicons are capable of expressing glycosylated proteins. The human respiratory syncytial virus (RSV) fusion (F) protein was chosen as a model glycoprotein and the HRV replicon DeltaP1FVP3 was generated by inserting the F protein-coding sequence in frame and in lieu of the 5' proximal 1489 nucleotides of the capsid-coding segment in the HRV-14 genome. When transfected into H1-HeLa cells, DeltaP1FVP3 replicated and led to the expression of the F protein. Inhibition with guanidine demonstrated that F-protein expression was dependent on DeltaP1FVP3 replication and did not result from translation of input RNA. Although most of the F protein remained as an immature, glycosylated precursor (F0), a readily detectable fraction of the protein was processed into the mature glycosylated subunit F1, an event known to occur within the Golgi apparatus. Packaged DeltaP1FVP3 replicons were generated in transfected HeLa cells by coexpression of homologous HRV capsid proteins using the vaccinia virus/T7 RNA polymerase hybrid system. Packaged replicon RNAs were capable of infecting fresh cells, leading to accumulation of the F protein as in RNA-transfected cells. Mice immunized with HeLa cell lysates containing F protein expressed from DeltaP1FVP3 produced neutralizing antibodies against RSV. These results indicate that an HRV-14 replicon can express a foreign glycosylated protein, providing further support for the potential of HRV replicons as a vaccine delivery system.

Animals↗
PubMed2001

Role of phospholipid transfer protein in rabbit lung development.

A 36-kDa phospholipid transfer protein (PLT-P(R)), which preferentially transfers phosphatidyl choline (PC) compared to phosphatidyl inositol (PI), was purified 827-fold from rabbit lung homogenate. Incorporation of cholesterol in unilamellar vesicles reduced the PC transfer activity of PLTP(R). Dipalmitoyl phosphatidyl choline uptake by alveolar type II cells was increased in the presence of the protein, and further enhanced in the presence of surfactant liposomes. However, a decrease in uptake was noted with cholesterol in host membranes. Incorporation of PI into host membranes had a low stimulatory effect on the process. All these effects were more pronounced in adult type II cells compared to premature, term and 3-day-old pups.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed2001

Alpha-synuclein has an altered conformation and shows a tight intermolecular interaction with ubiquitin in Lewy bodies.

Alpha-synuclein, a protein in which two mutations have been identified that cause autosomal dominant Parkinson's disease, is thought to serve as a nidus for the development of a Lewy body. We hypothesized that alpha-synuclein would display different intra- and intermolecular associations in Lewy bodies than it does in its normal intracellular compartments. Using sensitive fluorescence resonance energy transfer (FRET) techniques, we found evidence that alpha-synuclein is more compact and in closer association with other alpha-synuclein molecules in Lewy bodies than it is in the neuropil. In addition, we found evidence of a close, direct intermolecular interaction between the N terminus of alpha-synuclein and ubiquitin. These observations provide support for the hypothesis that in Lewy bodies alpha-synuclein adopts an altered three-dimensional structure and undergoes N-terminal ubiquitination.

Hippocampus↗
PubMed2001

Microkeratome-induced reduction of astigmatism after penetrating keratoplasty.

PURPOSE: To report the reduction in postpenetrating keratoplasty astigmatism with the use of the microkeratome to create a lamellar corneal flap as the first stage in a two-step laser in situ keratomileusis. METHODS: The hansatome microkeratome was used to create a lamellar corneal flap in a 24-year-old man with a net corneal astigmatism of 7.3 diopters, 2 years after penetrating keratoplasty. No laser ablation was performed. RESULTS: The net corneal astigmatism reduced to 3.9 diopters at 1 month and 2.3 diopters at 3 months of follow-up, without any laser ablation. CONCLUSION: Laser in situ keratomileusis may be performed as a two-stage procedure, because the lamellar corneal flap alone may reduce postpenetrating keratoplasty astigmatism.

Adult↗
PubMed2001

A close association of torsinA and alpha-synuclein in Lewy bodies: a fluorescence resonance energy transfer study.

TorsinA, a novel protein in which a mutation causes dominant, early onset torsion dystonia, may serve as a chaperone for misfolded proteins that require refolding or degradation. It has been hypothesized that misfolded alpha-synuclein, a protein in which two mutations cause autosomal dominantly inherited Parkinson's disease, serves as a nidus for the development of a Lewy body. We hypothesized that torsinA plays a role in the cellular processing of alpha-synuclein. We demonstrate that anti-torsin antibodies stain Lewy bodies and Lewy neurites in the substantia nigra and cortex. Using sensitive fluorescent resonance energy transfer (FRET) techniques, we find evidence of a close association between torsinA and alpha-synuclein in Lewy bodies.

Carrier Proteins↗
PubMed2001

Purification and characterization of dihydroxyacetone phosphate reductase from immature seeds of Brassica campestris L.

Dihydroxyacetone phosphate reductase (DHAP reductase) was purified to apparent electrophoretic homogeneity with about 24% recovery from immature seeds of Brassica campestris using (NH(4))(2)SO(4) fractionation, affinity chromatography, gel filtration and adsorption chromatography. The purified enzyme with molecular mass of about 62 kDa was a dimer with subunit molecular mass of 32 kDa. The enzyme exhibited maximum activity at pH 7.5 and was highly specific for NADH and DHAP. Typical Michaelis-Menten kinetics was obtained for both the substrates with K(m) values of 3.3 and 26.6 &mgr;M for NADH and DHAP, respectively. The enzyme did not require any metal ion for its activity. Rather, the activity was inhibited by Na(+), K(+), Mn(2+), Mg(2+,) and Ca(2+). ATP and fructose-1,6-P(2) inhibited the enzyme non-competitively with respect to DHAP with K(i) values of 0.96 and 1.3 mM, respectively. Substrate interaction kinetics and product inhibition studies were consistent with compulsory-ordered bi-bi reaction mechanism with NADH being the first substrate to bind and NAD being the last product to dissociate. Based on the properties discussed here, it appears that the enzyme probably functions for the production of glycerol-3-P from DHAP.

Journal Article↗
PubMed2001

Antifertility investigation and toxicological screening of the petroleum ether extract of the leaves of Mentha arvensis L. in male albino mice.

In male albino mice, the petroleum ether extract of the leaves of Mentha arvensis L., at the doses 10 and 20 mg/mouse per day for 20, 40 and 60 days, when administered orally, showed a dose and duration dependent reduction in the number of offspring of the treated male mated with normal females. Negative fertility was observed in both dose regimens after 60 days of treatment. The body weight and libido of the treated ammals remain unaffected. However, a significant decrease in the weight of the testis, epididymis, cauda epididymal sperm count, motility, viability and normal morphology of the spermatozoa was observed. The levels of serum protein, bilirubin, GOT, GPT and acid phosphatase, blood urea and haematological indices were unaltered throughout the course of investigation. All the altered parameters were reversible following withdrawal of treatment. The results suggest that the petroleum ether extract of the leaves of M. arvensis possess reversible antifertility property without adverse toxicity in male mice.

Alkanes↗
PubMed2001

Using nurse practitioners for skin cancer screening: a pilot study.

BACKGROUND: Skin cancer screening has the potential to detect early precancerous lesions and may ultimately be important in reducing melanoma mortality. The purpose of this study was to evaluate the ability of trained nurse practitioners to accurately identify suspicious lesions in a clinical setting. METHODS: We identified five nurse practitioners who had no previous experience in evaluating skin lesions. Each nurse practitioner participated in a training program for skin cancer detection consisting of a workshop, clinical apprenticeship, and didactic lectures. RESULTS: Evaluation of nurse practitioner competency involved three assessments. First, the nurse practitioner's ability to distinguish benign and malignant lesions was assessed using clinical color slides. The sensitivity of all five nurse practitioners to refer benign and malignant lesions for dermatologic follow-up based on the slides was 100%, whereas the specificity ranged from 53% to 100%. Second, each nurse practitioner evaluated approximately 25 different patients along with a single dermatologist. The nurse practitioner's ability to correctly refer patients with suspicious lesions for dermatologic follow-up was determined based on the dermatologist's assessment of need for referral. Results suggested a referral sensitivity and specificity ranging from 67% to 100% and 62% to 100%, respectively. In the final clinical assessment, 30 patients were independently examined by two dermatologists and four nurse practitioners. Using the consensus clinical diagnosis of the dermatologists as the gold standard, the nurse practitioner's sensitivity for detecting significant skin cancer lesions ranged from 50% to 100% and the detection specificity was 99% to 100%. CONCLUSIONS: These preliminary results have important implications for skin cancer screening efforts and suggest that nurse practitioners can be trained to accurately identify and triage suspicious lesions.

Clinical Competence↗
PubMed2001

Phacoemulsification combined with silicone oil removal through a posterior capsulorhexis.

PURPOSE: To evaluate the technique of silicone oil removal through a posterior capsulorhexis combined with phacoemulsification and intraocular lens (IOL) implantation. SETTING: Dr. Rajendra Prasad Center for Ophthalmic Sciences, New Delhi, India. METHODS: Fifteen eyes of 15 patients had phacoemulsification with removal of silicone oil, which had been used for intraocular tamponade after a previous pars plana vitrectomy. Eyes with a stable retina were included in the series. In all eyes, the silicone oil was removed through a planned posterior capsulorhexis after phacoemulsification. The parameters evaluated were the primary diagnosis, duration between silicone oil instillation and phacoemulsification, type of cataract, preoperative and postoperative best corrected visual acuities (BCVAs), and complications such as frequency of retinal redetachment and secondary cataract. RESULTS: Vitreoretinal surgery with silicone oil instillation was performed for rhegmatogenous-tractional detachment resulting from Eales' disease in 6 eyes and from proliferative diabetic retinopathy in 2 eyes, for primary rhegmatogenous retinal detachment in 6 eyes, and for traumatic rhegmatogenous detachment in 1 eye. The mean duration between the silicone oil instillation and phacoemulsification was 7.5 months +/- 3.8 (SD). Fourteen eyes had posterior subcapsular cataract, and 10 had nuclear sclerosis. Preoperative BCVA was worse than 6/60 in all eyes. The BCVA was 6/60 or better in 9 eyes after a minimum follow-up of 6 months. Two eyes had choroidal detachment in the early postoperative period. No eye had vitreous hemorrhage, retinal redetachment, secondary cataract, clinically significant endothelial decompensation or macular edema, or a dislocated IOL. CONCLUSION: The results indicate that silicone oil removal through a posterior capsulorhexis during phacoemulsification is a viable option and can be performed in selected cases of cataract with previous silicone oil instillation and a stable retina.

Adolescent↗
PubMed2001

Bimanual Sinskey hook technique to enlarge a preexisting capsulorhexis.

We describe a technique of enlarging a small primary capsulorhexis. Two Sinskey hooks are introduced through the side ports. One hook is placed above the anterior capsulorhexis margin with its tip pointing down, and the other is placed beneath the anterior capsulorhexis margin with its tip pointing up. A small nick is made between the tips of the 2 hooks, and the capsulorhexis is extended with a Utrata capsulorhexis forceps. This technique does not require additional instrumentation, impart zonular stress, or disturb wound architecture or integrity.

Capsulorhexis↗