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Biomedical subjects

P K Pandey

Publications and source records attributed to P K Pandey.

At least 19 recordsLinked to original sources
PubMed2006

Simultaneous intraocular and bilateral extraocular muscle involvement in a case of disseminated cysticercosis.

We report an unusual case of disseminated cysticercosis having simultaneous intraocular and extraocular involvement. This 21-year-old male presented with recent loss of vision in one eye and history of a single seizure. Examination revealed intravitreal cysticercosis with exudative retinal detachment. Computed tomographic scan showed disseminated neurocysticercosis and cysticercosis of the medial and inferior recti of both the eyes. The patient was started on albendazole and prednisolone under neurologist's supervision. Subsequent computed tomographic scan showed resolution of the extraocular muscle cysts and decrease in the number of neurocysticerci. The patient has had no further seizures. This is the first report of simultaneous intraocular and extraocular cysticercosis.

Adult↗
PubMed2006

Arsenic contamination in the Kanker district of central-east India: geology and health effects.

This paper identifies newer areas of arsenic contamination in the District Kanker, which adjoins the District Rajnandgaon where high contamination has been reported earlier. A correlation with the mobile phase episodes of arsenic contamination has been identified, which further hinges on the complex geology of the area. Arsenic concentrations in both surface and groundwater, aquatic organisms (snail and water weeds) soil and vegetation of Kanker district and its adjoining area have been reported here. The region has been found to contain an elevated level of arsenic. All segments of the ecoysystem are contaminated with arsenic at varying degrees. The levels of arsenic vary constantly depending on the season and location. An analysis of groundwater from 89 locations in the Kanker district has shown high values of arsenic, iron and manganese (mean: 144, 914 and 371 microg L(-1), respectively). The surface water of the region shows elevated levels of arsenic, which is influenced by the geological mineralised zonation. The most prevalent species in the groundwater is As(III), whereas the surface water of the rivers shows a significant contamination with the As(V) species. The analysis shows a bio-concentration of the toxic metals arsenic, nickel, copper and chromium. Higher arsenic concentrations (groundwater concentrations greater than 50 microg L(-1)) are associated with sedimentary deposits derived from volcanic rocks, hence mineral leaching appears to be the source of arsenic contamination. Higher levels of arsenic and manganese in the Kanker district have been found to cause impacts on the flora and fauna. A case study of episodic arsenical diarrhoea is presented.

Adolescent↗
PubMed2005

Cloning, restriction mapping and phylogenetic relationship of genomic components of MYMIV from Lablab purpureus.

The present work describes cloning of genomic components of whitefly transmitted geminivirus infecting Lablab purpureus syn. Dolichos lablab (commonly known as Dolichos bean or Hyacinth bean). The genome characterization using PCR with geminiviral degenerate primers and DNA sequencing were used to describe the bipartite virus associated with yellow mosaic disease of Dolichos lablab. Full-length DNA-A and DNA-B clones were obtained. The DNA-A sequence analysis showed that the isolate was similar to other Mungbean yellow mosaic India virus (MYMIV) isolates reported earlier. The nucleotide sequence analysis of the full-length DNA-A of virus isolate revealed more than 97% homology with Mungbean yellow mosaic India virus-[Cowpea] (AF481865), while the DNA-B also showed >95% homology with MYMIV-[Cp] (AF503580) and MYMIV-[Sb] (AY049771). The phylogenetic analysis of present isolate showed close relationship to legume geminiviruses. The nucleotide sequence analysis showed presence of six open reading frames (ORFs) in DNA-A, with 2 ORFs aligned in sense and 4 ORFs in antisense orientation. Similarly, DNA-B contained two open reading frames (ORFs), one in sense and another in antisense orientation.

Cloning, Molecular↗
PubMed2005

Removal of arsenic(III) from aqueous solutions using fresh and immobilized plant biomass.

The ability of Garcinia cambogia, an indigenous plant found in many parts of India, to remove trivalent arsenic from solution was assessed. Batch experiments were carried out to characterize the As(III) removal capability of fresh and immobilized biomass of G. cambogia. It was found that the kinetic property and uptake capacity of fresh biomass were significantly enhanced by the immobilization procedure. The uptake of As(III) by fresh and immobilized biomass was not greatly affected by solution pH with optimal biosorption occurring at around pH 6--8. The presence of common ions such as Ca and Mg at concentrations up to 100mg/l had no effect on As(III) removal. However, the presence of Fe(III) at 100mg/l caused a noticeable drop in the extent of As(III) removal but the effect was minimal when Fe(III) was present at 10mg/l. The adsorption isotherms quantitatively predicted the extent of As(III) removal in groundwater samples collected from an arsenic-contaminated site in India. Immobilized biomass loaded with As(III) was amenable to efficient regeneration with NaOH solution. Column studies showed that immobilized biomass could be reused over five cycles of loading and elution. The excellent As(III) sequestering capability of fresh and immobilized G. cambogia biomass could lead to the development of a viable and cost-effective technology for arsenic removal in groundwater.

Adsorption↗
PubMed2002

Physiological and biochemical alterations in Anabaena 7120 under iron stress.

Various physiological and biochemical process like growth, NO3- -uptake, nitrate reductase, glutamine synthetase and ATPases (Mg2+ and Ca2+ dependent) in the cyanobacterium Anabaena 7120 were observed under iron stress. Growth was found to be maximum in 50 microM Fe3+ added cells however, 20 microM Fe3+ (the Fe3+ concentration generally used for routine culturing of cyanobacterial cell in Chu 10 medium) incubation resulted in lower growth. Fe3+ starvation on the other hand showed very poor growth up to 4th day but once the growth started it reached at significant level on 7th day. Higher Fe3+ concentration reflected reduced growth with lethality at 500 microM Fe3+. Chlorophyll a fluorescence under Fe3+ stress reflected almost the similar results as in case of growth. However, the pigment was found to be more sensitive as compared to protein under Fe3+ stress. Similar results have been observed in case of NO3-uptake with only 80% reduction in nutrient uptake in 500 microM Fe3+ incubated cells. Nitrate reductase activity was lower in Fe3+ starved cells as compared to significant enzyme activity in 20 and 50 microM Fe3+ incubated cells. Similar to nitrate reductase, glutamine synthetase also showed maximum level in 50 microM Fe3+ added cells, however, higher Fe3+ concentration (300-500 microM ) resulted in reduced enzymatic activity. Glutamine synthetase activity was less sensitivity as compared to nitrate reductase activity under Fe3+ stress. ATPase (Mg2+ and Ca2+ dependent) always showed higher level with increasing Fe3+ concentration.

Adenosine Triphosphatases↗
PubMed2001

The beta7-beta8 loop of the p51 subunit in the heterodimeric (p66/p51) human immunodeficiency virus type 1 reverse transcriptase is essential for the catalytic function of the p66 subunit.

The heterodimeric human immunodeficiency virus type 1 reverse transcriptase (HIV-1 RT) is composed of p66 and p51 subunits, p66 being the catalytic subunit. Our earlier investigation on the role of p51 in the catalytic process has shown that the p51 subunit facilitates the loading of the p66 subunit onto the template primer (TP). We had postulated that the beta7-beta8 loop of the p51 subunit may be involved in opening the polymerase cleft of p66 for DNA binding [Pandey, V. N., et al. (1996) Biochemistry 35, 2168]. We report here that deletion or alanine substitution of four residues of the beta7-beta8 loop results in severe impairment of the polymerase function of the heterodimeric enzyme. The enzyme activity was restored to the wild-type levels when the mutant p66 subunit was dimerized with the wild-type p51, suggesting that the intact beta7-beta8 loop in the p51 subunit is indispensable for the catalytic function of p66. Further, the template primer binding ability of the enzyme was significantly reduced upon deletion or alanine substitution in the beta7-beta8 loop. Interestingly, the loss of the TP binding ability of the mutant p66 was restored upon dimerization with wild-type p51. Examination of the glycerol gradient ultracentrifugation analysis revealed that while the wild-type HIV-1 RT sediments as a dimeric protein, the mutant enzymes carrying deletion or alanine substitution in both the subunits sediment predominantly as monomeric proteins, suggesting their inability to form stable dimers. In contrast, mutant p66 dimerized with wild-type p51 (p66delta/p51WT and p66Ala/p51WT) sedimented at the dimeric position. Taken together, these results clearly implicate the importance of the beta7-beta8 loop of p51 in the formation of stable functional heterodimers.

Amino Acid Substitution↗
PubMed2001

Extraocular muscle cysticercosis presenting as Brown syndrome.

PURPOSE: Report of a case of acquired Brown syndrome caused by infestation of the superior oblique muscle by Cysticercus cellulosae. METHOD: Case seen in a referral practice. A 20-year-old man presented with recurrent attacks of conjunctivitis and diplopia in upgaze. Clinical examination of ocular motility established a diagnosis of acquired Brown syndrome of the right eye. Computed tomography of the right orbit unequivocally established the diagnosis of superior oblique muscle cysticercosis. The patient was started on systemic steroids and albendazole in the prescribed doses for a month. RESULT: Serial computed tomography scans of the orbit revealed resolution of the cystic lesion after a month. Clinically, although there was restoration of ocular motility in upgaze, mild restriction of movement of the right eye in levoelevation persisted. However, the patient was symptomatically better with amelioration of the recurrent conjunctivitis and diplopia in primary gaze. CONCLUSION: Extraocular muscle cysticercosis should be considered in the differential diagnosis of acquired motility disorder. The presentation of extraocular muscle cysticercosis as an acquired Brown syndrome is unusual. Response to medical therapy was satisfactory.

Adult↗
PubMed2001

Insertion of a peptide from MuLV RT into the connection subdomain of HIV-1 RT results in a functionally active chimeric enzyme in monomeric conformation.

The natural form of the human immunodeficiency virus type one reverse transcriptase (HIV-1 RT) found in virion particles is a heterodimer composed of the p66 and p51 subunits. The catalytic activity resides in the larger subunit in the heterodimeric (p66/p51) enzyme while in the monomeric form it is inactive. In contrast, Murine leukemia virus RT (MuLV RT) is functionally active in the monomeric form. In the primary amino acid sequence alignment of MuLV RT and HIV-1 RT, we have identified three specific regions in MuLV RT, that were missing in HIV-1 RT. In a separate study, we have shown that a chimeric RT construct comprising of the polymerase domain of HIV-1 RT and RNase-H domain of MuLV RT is functionally active as monomer [20]. In this communication, we demonstrate that insertion of a peptide (corresponding to amino acid residues 480-506) from the connection subdomain of MuLV RT into the connection subdomain of HIV-1 RT (between residues 429 and 430) results in a functionally active monomeric chimeric RT. Furthermore, this chimeric enzyme does not dimerize with exogenously added p51 subunit of HIV-1RT. Functional analysis of the chimeric RT revealed template specific variations in its catalytic activity. The chimeric enzyme catalyzes DNA synthesis on both heteropolymeric DNA and homopolymeric RNA (poly rA) template but curiously lacks reverse transcriptase ability on heteropolymeric RNA template. Similar to MuLV RT, the polymerase activity of the chimeric enzyme is not affected by acetonitrile, a reagent which dissociates dimeric HIV-1 RT into inactive monomers. These results together with a proposed 3-D molecular model of the chimeric enzyme suggests that the insertion of the missing region may induce a change in the spatial position of RNase H domain such that it is functionally active in monomeric conformation.

Acetonitriles↗
PubMed2000

Inhibition of Tat-mediated transactivation of HIV-1 LTR transcription by polyamide nucleic acid targeted to TAR hairpin element.

Tat, an essential human immunodeficiency virus type 1 protein interacts with the transactivation response element (TAR) and stimulates transcription from the viral long-terminal repeat (LTR). Blockage of Tat-TAR interaction halts viral transcription and hence replication. We have found that polyamide nucleic acid (PNA), targeted to the TAR sequences of viral RNA genome is able to prevent Tat-TAR interaction by efficient sequestration of the TAR. Anti-TAR PNA competes for TAR and prevents Tat-mediated stimulation of HIV-1 LTR transcription in vitro but has no influence on the basal level of transcription in the absence of Tat. Using a reporter gene construct pHIV LTR-CAT and pCMV-Tat in cell culture, we have further shown that anti-TAR PNA is able to block Tat-mediated transactivation of HIV-1 LTR transcription in vivo as judged by the extent of LTR driven CAT gene expression in the absence and presence of anti-TAR PNA. Supplementation of 100 nM of anti-TAR PNA into the culture medium further enhances the suppression of transactivation. Nonspecific scrambled PNA had no influence on Tat-TAR interaction and LTR-driven CAT gene expression in cell culture. These results suggest that PNA targeted to the TAR sequence of the viral genome may be a potential inhibitor of HIV-1 gene expression.

Antiviral Agents↗
PubMed2000

Role of glutamine 151 of human immunodeficiency virus type-1 reverse transcriptase in substrate selection as assessed by site-directed mutagenesis.

A natural mutation at codon 151 (Gln --> Met; Q151M) of HIV-1 RT has been shown to confer resistance to the virus against dideoxy nucleoside analogues [Shirasaka, T., et al. (1995) Proc. Natl. Acad. Sci. U.S.A. 92, 2398], suggesting that Gln 151 may be involved in conferring sensitivity to nucleoside analogues. To understand its functional implication, we generated two mutant derivatives of this residue (Q151M and Q151N) and examined their sensitivities to ddNTPs and their ability to discriminate against rNTPs versus dNTP substrates on natural U5-PBS HIV-1 RNA template. We found that Q151M was highly discriminatory against all four ddNTPs but was able to incorporate rNTPs as efficiently as the wild type enzyme. In contrast, the Q151N mutant was only moderately resistant to ddNTPs but exhibited a higher level of discrimination against rNTPs. The fidelity of misinsertion was found to be highest for the Q151N mutant followed by Q151M and the wild type enzyme. These results point toward the importance of the amino acid side chain at position 151 in influencing the ability of the enzyme in recognition and discrimination against the sugar moieties of nucleotide substrates.

Asparagine↗
PubMed1998

Functional analysis of amino acid residues constituting the dNTP binding pocket of HIV-1 reverse transcriptase.

In order to understand the functional implication of residues constituting the dNTP-binding pocket of human immunodeficiency virus type 1 reverse transcriptase, we performed site-directed mutagenesis at positions 65, 72, 113, 115, 151, 183, 184, and 219, and the resulting mutant enzymes were examined for their biochemical properties and nucleotide selectivity on RNA and DNA templates. Mutations at positions 65, 115, 183, 184, and 219 had negligible to moderate influence on the polymerase activity, while Ala substitution at positions 72 and 151 as well as substitution with Ala or Glu at position 113 severely impaired the polymerase function of the enzyme. The K219A, Y115F, and Q151M mutants had no influence on the fidelity; Y183A, Y183F, K65A, and Q151N mutants exhibited higher fidelity on both RNA and DNA templates, while Y115A was less error-prone selectively on a DNA template. Analysis of the three-dimensional model of the enzyme-template primer-dNTP ternary complex suggests that residues Tyr-183, Lys-65, and Gln-151 may have impact on the flexibility of the dNTP-binding pocket by virtue of their multiple interactions with the dNTP, template, primer, and other neighboring residues constituting the pocket. Recruitment of the correct versus incorrect nucleotides may be a function of the flexibility of this pocket. A relatively rigid pocket would provide greater stringency, resulting in higher fidelity of DNA synthesis in contrast to a flexible pocket. Substitution of a residue having multiple interactions with a residue having reduced interaction capability will alter the internal geometry of the pocket, thus directly influencing the fidelity.

Amino Acids↗
PubMed1998

The p51 subunit of human immunodeficiency virus type 1 reverse transcriptase is essential in loading the p66 subunit on the template primer.

Human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) is a dimeric enzyme consisting of p66 and p51 subunits. The functional role of the p51 subunit remains elusive since all the catalytic functions appear to be executed through the p66 subunit. We report here that the p51 subunit, in addition to providing structural support to the p66 subunit, may be involved in facilitating the loading of the p66 subunit on to the template-primer (TP). This possibility is supported by following observations: (i) Upon binding to the TP, the p51 subunit can be dissociated by acetonitrile treatment and the template-primer-bound p66 monomer alone is capable of catalyzing DNA synthesis. (ii) Photo-cross-linking of template-primer to HIV-1 RT is abolished by dissociation of the p51 subunit prior to the TP binding but remains unaffected after the TP binding step. (iii) The p66-TP covalent complex selectively generated by UV irradiation and separated by gel electrophoresis can incorporate a single nucleotide in situ upon its renaturation in the gel. (iv) Treatment of HIV-1 RT with (tert-butyldimethylsilyl)spiroaminooxathioledioside (TSAO), an inhibitor that specifically binds to the beta7 beta8 loop of p51, destabilizes the heterodimeric enzyme, resulting in the subsequent loss of DNA binding.

Acetonitriles↗
PubMed1998

An enzymatically active chimeric HIV-1 reverse transcriptase (RT) with the RNase-H domain of murine leukemia virus RT exists as a monomer.

The existence of retroviral reverse transcriptases as monomers or dimers is rather intriguing. A classical example of the former is murine leukemia virus reverse transcriptase (MuLV RT), while human immunodeficiency virus type 1 (HIV-1) RT represents the latter. A careful scrutiny of the amino acid sequence alignment of the two enzymes pinpoints the region tentatively responsible for this phenomenon. We report here the construction of a chimeric enzyme containing the first 425 amino acid residues from the N-terminal domain of HIV-1 RT and 200 amino acid residues from the C-terminal domain of MuLV RT. The chimeric enzyme exists as a monomer with intact DNA polymerase and RNase-H functions.

Amino Acid Sequence↗
PubMed1998

Polyamide nucleic acid-DNA chimera lacking the phosphate backbone are novel primers for polymerase reaction catalyzed by DNA polymerases.

A peptide nucleic acid (PNA) oligomer, an analogue of DNA, was examined for its ability to function as a primer or a template to support DNA synthesis catalyzed by DNA polymerases. The analogue, (PNA)19-TpG-OH, comprised of 19 bases in the form of PNA followed by a dinucleotide (TpG-OH) with a single phosphate and a free 3'OH terminus, was recognized as a bona fide primer by 2 reverse transcriptases and also by the Klenow fragment of E. coli DNA polymerase I. The 21-mer PNA chimera is extended on both RNA and DNA templates by all three polymerases. The specificity of binding of the PNA chimeric primer/DNA template at the template-primer binding site of the enzyme was shown by its photo-cross-linking ability to the enzyme which could be effectively competed out by another TP but not by template or primer alone. Furthermore, the chimeric TP-enzyme covalent complex was found to be catalytically active as judged by its ability to incorporate one nucleotide onto the 3'OH terminus of the immobilized primer. PNA sequences were also recognized as template when annealed with a DNA primer. These observations are in variance with the conventional suggestion that the phosphate backbone in the duplex region is essential for recognition and binding by DNA polymerases. The efficient extension of (PNA)19-TpG-OH suggests that the diameter of the duplex region of template primer rather than the phosphate backbone may be sufficient for recognition by DNA polymerases.

Binding Sites↗
PubMed1998

Calcium-dependent metabolic regulations in prokaryotes indicate conserved nature of calmodulin gene.

Role of free calcium and calcium binding protein calmodulin as signal molecule in cellular regulation is well established in eukaryotes. However, reports on Ca(2+)-dependent processes and their inhibition by calcium and/or calmodulin antagonists indicate towards the presence of calmodulin in prokaryotes as well. The common evolutionary origin of pro- and eukaryotes and many examples of evolutionary conservation of structure and functions support the contention of such conservation of the role of Ca2+ and calmodulin. Eukaryotic calmodulin (CaM) contains four structurally and functionally similar Ca2+ domains named I, II, III and IV. Each Ca2+ binding loop consists of 12 amino acid residues with ligands arranged spatially to satisfy the octahedral symmetry of Ca2+ binding. Plant calmodulin differ from vertebrate ones in 13 to 14 amino acid positions of which nine occur at -COOH- terminal half. Differences between protozoan and mammalian CaM also occur mostly in the same half. Isolation and characterization, although to a little extent, of CaM-like proteins from bacteria and cyanobacteria and their comparison with CaMs from diverse origin suggest high degree of conservation. Non-bulky amino acids like glycine, alanine and serine with low specific rotation are present in greater number in the primitive form of calmodulin and have been significantly reduced in highly evolved form of calmodulin, suggesting that their requirement was insignificant and were eliminated from EF hand structure during evolution. However, amino acids like glutamate/glutamine and aspartate/asparagine were highly conserved and did not show any major change in their frequency since their positions are too significant in calcium binding domain. While the number of positively charged amino acids like arginine and leucine was increased, histidine containing weakly ionized group and having a significant buffering capacity was reduced to a major extent, further suggesting that the acidic nature of calmodulin protein has been maintained during evolution. Thus it is now clear that the entire superfamily of Ca2+ binding proteins have arisen from a common genetic ancestry. Two successive tandem duplications of gene encoding a single domain containing protein of 30-40 residues gave rise to a four domain molecule from which this family was then derived.

Amino Acid Sequence↗
PubMed1997

Short-term results of initial trabeculectomy with intraoperative or postoperative 5-fluorouracil for primary glaucomas.

Thirty-three eyes of 33 patients were prospectively evaluated to study the short term efficacy, safety, and overall surgical outcome of initial trabeculectomy for primary glaucomas with adjunctive intraoperative or postoperative 5-Fluorouracil (5-FU) use. Twelve eyes serving as controls underwent trabeculectomy without adjunctive antimetabolites. Eleven eyes received intraoperative 5-FU, while 10 eyes received subconjunctival 5-FU postoperatively. Intraocular pressure (IOP) was maintained below 22 mmHg at 3 months of follow up in 90.9% and 80% of patients in the intraoperative and postoperative 5-FU groups, respectively, without use of additional antiglaucoma medications, whereas 66.7% of the patients in the control group achieved similar IOP levels. Hypotony (IOP < 6 mmHg) was seen more commonly after intraoperative 5-FU (27.3%). Corneal epithelial defects were seen exclusively in the postoperative 5-FU group (40%). The use of intraoperative 5-FU exclusively as a mode of antimetabolites delivery seems an acceptable alternative to enhance success rates of trabeculectomy for the primary glaucomas.

Administration, Topical↗