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R Bhatt

Publications and source records attributed to R Bhatt.

48 records · Page 3Linked to original sources
PubMed1987

Structure-function analysis of human interleukin-2. Identification of amino acid residues required for biological activity.

To locate functional domains of the interleukin-2 (IL-2) protein, a cDNA clone encoding biologically active human IL-2 was mutagenized using synthetic oligonucleotides to incorporate defined amino acid substitutions and deletions in the mature protein. The IL-2 analogs were then produced in Escherichia coli and assayed for the ability to induce proliferation of IL-2-dependent cells and the ability to compete for binding to the IL-2 receptor. Our analysis of over 50 different mutations demonstrated that the integrity of at least three regions of the IL-2 molecule is required for full biological activity: the NH2 terminus (residues 1-20), the COOH terminus (residues 121-133), and 2 of the 3 cysteine residues (58 and 105). Deletion of the NH2-terminal 20 amino acids or the COOH-terminal 10 amino acids resulted in the loss of greater than 99% of bioactivity and binding. Amino acid substitutions at specific positions in these regions also resulted in proteins which retained less than 1% activity. The NH2 terminus and an adjacent internal region were recognized by neutralizing anti-IL-2 antibodies. In combination with the results from epitope competition analysis with neutralizing antibodies, these data are consistent with the IL-2 protein being folded such that the NH2 terminus, the COOH terminus, and the internal 30- to 60-region are juxtaposed to form the binding site recognized by the IL-2 receptor.

Amino Acid Sequence↗
PubMed1985

Conformation and dynamics of the Pribnow box region of the self-complementary d(C-G-A-T-T-A-T-A-A-T-C-G) duplex in solution.

Nuclear magnetic resonance (NMR) has been used to monitor the conformation and dynamics of the d(C1-G2-A3-T4-T5-A6-T6-A5-A4-T3-C2-G1) self-complementary dodecanucleotide duplex (henceforth called Pribnow 12-mer), which contains a TATAAT Pribnow box and a central core of eight dA X dT base pairs. The exchangeable imino and nonexchangeable base protons have been assigned from one-dimensional intra and inter base pair nuclear Overhauser effect (NOE) measurements. Premelting conformational changes are observed at all the dA X dT base pairs in the central octanucleotide core in the Pribnow 12-mer duplex with the duplex to strand transition occurring at 55 degrees C in 0.1 M phosphate solution. The magnitude of the NOE measurements between minor groove H-2 protons of adjacent adenosines demonstrates that the base pairs are propeller twisted with the same handedness as observed in the crystalline state. The thymidine imino proton hydrogen exchange at the dA X dT base pairs has been measured from saturation recovery measurements as a function of temperature. The exchange rates and activation barriers show small variations among the four different dA X dT base pairs in the Pribnow 12-mer duplex.(ABSTRACT TRUNCATED AT 250 WORDS)

Base Sequence↗
PubMed1985

Four-year follow-up of insertion of quinacrine hydrochloride pellets as a means of nonsurgical female sterilization.

Eighty-four women were admitted to a study in Baroda, India, designed for evaluation of the efficacy of three transcervical insertions of quinacrine hydrochloride pellets, each 1 month apart, in producing occlusion of the oviducts. A 4-year follow-up has been completed for 100% of the women. Three women became pregnant during the time between the first and third administrations. Of the 81 women remaining in the study after administrations were complete, 3 became pregnant during the 4-year follow-up period, which resulted in a cumulative life-table pregnancy rate of 3.7 at 48 months. The results of this study indicate that intrauterine insertion of quinacrine pellets can be a safe, effective nonsurgical sterilization procedure.

Adult↗
PubMed1984

Molecular cloning and sequence determination of rat preproenkephalin cDNA: sensitive probe for studying transcriptional changes in rat tissues.

A cDNA probe was prepared to investigate the regulation of proenkephalin biosynthesis in the rat. This was necessary because human and bovine proenkephalin cDNA were not sensitive enough for the accurate detection of preproenkephalin mRNA in tissues that contain low copy numbers of this message, such as the adrenal gland. The rat probe was prepared in the following manner. Preproenkephalin mRNA was enriched by sucrose gradient centrifugation of poly(A)-containing mRNA from rat brain and was used as a template for double-stranded cDNA synthesis. The resulting cDNA was inserted into the plasmid pBR322, and recombinant plasmids were used to transform Escherichia coli RR1 cells. A synthetic oligodeoxyribonucleotide (30 bases long) with a sequence that had previously been shown to be identical in bovine and human preproenkephalin cDNA was prepared to screen the clone bank. The plasmid with the longest cDNA insert (about 1200 bases) from the positive clones was isolated, and the sequence of the entire protein coding region was determined. Like the bovine and human gene products, rat preproenkephalin contains four [Met]enkephalin sequences and one copy each of [Leu]enkephalin, [Met]enkephalin-Arg6-Gly7-Leu8, and [Met]enkephalin-Arg6-Phe7. Rat preproenkephalin is 80% and 83% homologous to the bovine and human forms, respectively, at the nucleotide level and is 82% homologous to both species at the amino acid level. Rat preproenkephalin contains 269 amino acid residues, making it larger than the human (267 residues) and bovine (263 residues) precursors. The sensitivity for detection of rat preproenkephalin mRNA with the rat cDNA was several times greater than with the corresponding cDNAs from bovine and human sources.

Amino Acid Sequence↗
PubMed1983

Sequence dependence of base-pair stacking in right-handed DNA in solution: proton nuclear Overhauser effect NMR measurements.

Single-crystal x-ray studies of d(C-G-C-G-A-A-T-T-C-G-C-G) exhibit base-pair propeller twisting [Dickerson, R. E. & Drew, H. R. (1981) J. Mol. Biol. 149, 761-786] that results in close contacts between adjacent purines in the minor groove in pyrimidine (3'-5')-purine steps and in the major groove in purine (3'-5')-pyrimidine steps [Calladine, C. R. (1982) J. Mol. Biol. 161, 343-362]. These observations require an approximately 3.4 A separation between the minor groove edges of adenosines on adjacent base pairs for the dA-dA step but predict a smaller separation for the dT-dA step and a larger separation for the dA-dT step in a D(A-T-T-A).d(T-A-A-T) fragment. We have confirmed these predictions from steady-state nuclear Overhauser effect measurements between assigned minor groove adenosine H-2 protons on adjacent base pairs in the proton NMR spectrum of the d(C1-G2-A3-T4-T5-A6-T6-A5-A4-T3-C2-G1) self-complementary dodecanucleotide duplex (henceforth called the Pribnow 12-mer) in solution. The measured cross-relaxation rates (product of steady-state nuclear Overhauser effect and selective spin- lattice relaxation rates) translate to interproton separations between adjacent adenosine H-2 protons of 4.22 A in the (dA3-dT4).(dA4-dT3) step, of 3.56 A in the (dT4-dT5).dA5-dA4) step, and of 3.17 A in the (dT5-dA6).(dT6-dA5) step for the Pribnow 12-mer duplex with an isotropic rotational correlation time of 9 ns at 5 degrees C. These proton NMR results show that the sequence-dependent base-pair stacking resulting from base-pair propeller twisting of defined handedness for right-handed DNA in the solid state is maintained in aqueous solution.

Base Composition↗
PubMed1981

A set of synthetic oligodeoxyribonucleotide primers for DNA sequencing in the plasmid vector pBR322.

Seven oligonucleotide primers complementary to the plasmid vector pBR322 at positions adjacent to five of the unique restriction endonuclease cleavage sites (EcoRI, HindIII, BamHI, SalI and PstI) have been chemically synthesized. The polarity of the primers is such that any DNA inserted at one or a combination of two of the above restriction sites may be sequenced by the chain termination method using one of the synthetic DNA primers. One of the primers for sequencing inserts at the PstI site of pBR322 is also complementary to the M13 phage vector designated bla6. This set of universal primers is useful for rapid sequence determination of DNA cloned into pBR322 or M13bla6.

Base Sequence↗
PubMedDate not supplied

Detection of nucleic acids by cycling probe technology on magnetic particles: high sensitivity and ease of separation.

Cycling Probe Technology (CPT) is a signal amplification system that allows detection of nucleic acid target sequences without target amplification. CPT employs a sequence specific chimeric probe, typically DNA-RNA-DNA, which hybridizes to a complementary target DNA sequence and becomes a substrate for RNase H. Cleavage occurs at the RNA internucleotide linkages and results in dissociation of the probe from the target, thereby making it available for the next probe molecule. This communication describes the use of oligonucleotides attached to solid supports for target capture and release followed by solution and solid phase cycling. Through the attachment of chimeric probes to Sera-Mag magnetic particles (SMP) a simple and effective method of separating the cleaved probe from non-cycled probe has been developed. By capturing the target DNA on particles and separating it from the extraneous non-specific DNA we are able to dramatically reduce background and thus discriminate between samples of Methicillin Resistant (MRSA) and Methicillin Sensitive (MSSA) Staphylococcus Aureus. We conjugated oligonucleotide probes to SMPs (approximately 1 um) and Nylon beads (NB) which were coated with ID Biomedical's proprietary coating materials (R, patent pending). The general structure of the constructs is shown below: [table: see text]

Magnetics↗
PubMedDate not supplied

Comparative pathogenicity of trichomonas vaginalis isolated from symptomatic & asymptomatic cases.

Pathogenicity of 19 isolates of T.vaginalis obtained from vaginal specimens were studied in the murine model by intraperitoneal route. Sixteen isolates were recovered from the females with various clinical conditions and 3 isolates were from normal healthy females. Pathogenicity level of these isolates were studied by inoculating 5 mice per isolates through intraperitoneal route and the animals were sacrificed on tenth day post-inoculation. In general, all the isolates recovered produced infection in mice. On comparison with the reference strain obtained from Hoechst India Ltd., seven isolates recovered from symptomatic cases and one strain from healthy females produced severe infection in mice. Though variation in pathogenicity level was observed among the isolates, a definite correlation between clinical picture in natural host and pathogenicity in mice was not observed.

Animals↗
PubMedDate not supplied

Detection of serum antitrichomonal antibodies in urogenital trichomoniasis by immunofluorescence.

Trichomonas vaginalis is a frequently encountered genital pathogen in both males and females. In females, vaginitis due to this parasite is one of the most common manifestation. The indirect fluorescent technique (IFA) test was carried out to detect antitrichomonal antibodies in 370 female patients using whole cell antigen. Seventy one (19.18%) gave positive reaction for either of the class IgG, IgM and IgA antibodies. The level of the IgG class antibodies was found to be higher i.e. 58 (81.69%) than IgM 11 (15.27%) antibodies, which may be suggestive of past infection or a prolonged manifestation by the organisms.

Antibodies, Anti-Idiotypic↗
PubMedDate not supplied

Comparative pathogenicity of Trichomonas vaginalis isolated from symptomatic and asymptomatic cases.

Pathogenicity of 19 isolates of Trichomonas vaginalis obtained from vaginal specimens were studied in the murine model by intraperitoneal inoculation. Sixteen isolates were recovered from the females with various clinical conditions and 3 isolates were from normal healthy females. Pathogenicity level of these isolates were studied by inoculating 5 mice per isolate through intraperitoneal route and the animals were sacrificed on the 10th day post-inoculation. In general, all the isolates recovered produced infection in mice. On comparison with the reference strain obtained from Hoechst India Ltd., seven isolates recovered from symptomatic cases and one strain from healthy females produced severe infection in mice. Though variation in the pathogenicity level was observed among the isolates, a definite correlation between the clinical picture in the natural host and pathogenicity in mice was not observed.

Animals↗