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Biomedical subjects

P Sarkar

Publications and source records attributed to P Sarkar.

At least 55 records · Page 3Linked to original sources
PubMed1979

Termination of transcription by Escherichia coli RNA polymerase: influence of secondary structure of RNA transcripts on rho-independent and rho-dependent termination.

The effect of RNA secondary structure on rho-independent and rho-dependent termination of transcription of T3 DNA by Escherichia coli RNA polymerase has been studied by incorporating, into nascent transcripts, base analogs that lead to altered base-pairing properties. A guanine --> hypoxanthine substitution, with attendant weakening of secondary structure, abolished the rho-independent termination at 20% of the genome; in contrast, replacement of cytosine with 5-bromocytosine, which forms stronger pairs with guanine, enhanced termination at this site. rho-Independent termination was not altered by replacing uracil with 5-bromouracil. There are two major rho-dependent termination sites on the T3 DNA-at 8 and 15%. The termination activity of rho in this system also depended on RNA secondary structure. The incorporation of 5-bromouracil instead of uracil into RNA did not alter the site specificity of rho action but rho was rendered inactive when cytosine was replaced by 5-bromocytosine. In contrast, replacement of GTP with ITP in the reaction increased rho-dependent inhibition of RNA synthesis, caused production of heterogeneous-sized transcripts, and stimulated rho-mediated ATP hydrolysis. The rho-associated ATPase activity, in the presence of isolated T3 RNA, was also stimulated by inosine substitution. Furthermore, the temperature-sensitive rho isolated from rho 15 mutant of E. coli, which does not terminate transcription in the presence of the common rNTPs, was active when GTP was replaced with ITP. These results suggest that strongly paired G.C-rich regions in RNA stem-loop structures or RNA.DNA hybrids are essential for rho-independent termination, whereas rho-dependent termination requires weakly paired cytosine residues for its action.

Adenosine Triphosphatases↗
PubMed1977

Function of initiation factor 1 in the binding and release of initiation factor 2 from ribosomal initiation complexes in Escherichia coli.

1. Studies on the function of initiation factor 1 (IF-1) in the formation of 30 S initiation complexes have been carried out. IF-1 appears to prevent the dissociation of initiation factor 2 (IF-2) from the 30 S initiation complex. The factor has no effect on either the initial binding of IF-2 nor does it increase the amount of IF-2 dependent fMet-tRNA and GTP bound to the 30 S subunit. Bound fMet-tRNA remains stable to sucrose gradient centrifugation even in the absence of IF-1. 2. It is postulated that the presence of IF-2 on the 30 S complex is necessary so that at the time of junction with the 50 S subunit to form a 70 S complex, the 70 S-dependent GTPase activity of IF-2 can hydrolyze GTP. This hydrolysis provides a means by which GTP can be removed to facilitate formation of a 70 S initiation complex active in peptidyl transfer. In support of this postulate, it was observed that 30 S initiation complexes formed in the absence of IF-1 could be depleted of their complexes were still able to accept 50 S subunits to form 70 S complexes which could still donate fMet-tRNA into peptide linkages. These results indicate that 30 S complexes lacking GTP do not require IF-2 for formation of active 70 S complexes. 3. IF-1, which is required to prevent dissociation of IF-2 from the 30 S initiation complex, is also required for release of IF-2 from ribosomes following 70 S initiation complex formation. The mechanisms of the release of IF-2 has been studied in greater detail. Evidence is presented which rules out the presence of a stable IF-2 GDP complex on the surface of the 70 S ribosome following GTP hydrolysis and of any exchange reactions between IF-1 and guanine nucleotides necessary for effecting the release of IF-2. IF-2 remains on the 70 S initiation complexes after release of guanine nucleotides and can be liberated solely by addition of IF-1.

Binding Sites↗
PubMed1974

Thiostrepton inhibition of initiation factor 1 activity in polypeptide chain initiation in Escherichia coli.

Thiostrepton, a peptide antibiotic, inhibits the GTP-dependent 70S initiation complex formation (as measured by binding of fMet-tRNA to ribosomes and concomitant hydrolysis of GTP) only when initiation factor 1 is present to permit catalytic recycling of initiation factor 2 in the initiation reaction. When initiation factor 1 is absent, the binding of fMet-tRNA and GTP hydrolysis occur stoichiometrically with respect to initiation factor 2, and thiostrepton has no effect on either reaction under these conditions. Detailed analysis of this inhibition process shows that thiostrepton prevents catalytic recycling of initiation factor 2 by blocking the action of initiation factor 1, which is required for the dissociation of initiation factor 2 from the 70S initiation complex. This dissociation is necessary for the catalytic reutilization of initiation factor 2 in the initiation reaction. The antibiotic does not directly inhibit GTP hydrolysis per se in initiation. The inhibition of fMet-tRNA binding to ribosomes by thiostrepton is also dependent on the concentration of GTP; the inhibition is most pronounced at low concentrations of GTP, but at a high molar ratio of GTP to thiostrepton, the inhibition is completely abolished.

Anti-Bacterial Agents↗
PubMed1972

Release of polypeptide chain initiation factor IF-2 during initiation complex formation.

Polypeptide chain initiation factor IF-2 binds to 30S ribosomal subunits. This binding is enhanced by IF-1 and IF-3. During GTP-dependent formation of a 70S initiation complex, IF-2 is released from the ribosome. During 70S initiation complex formation dependent on the methylene analogue of GTP, GMPPCH(2)P, IF-2 is not released, but remains bound to the 70S ribosome. This result suggests that IF-2 release requires GTP hydrolysis. In agreement with this presumed requirement, IF-2 functions catalytically with GTP, but stoichiometrically with GMPPCH(2)P, in bringing about 70S initiation complex formation.

Centrifugation, Density Gradient↗
PubMedDate not supplied

A sequential scanning of the immune efficiency in astrocytoma (Grade I to Grade Iii), meningioma and secondary glioma patients with and without therapeutic scheduling.

PURPOSE: Glioma induces immune suppression. However, data revealing the immune status in glioma patients with sequential therapeutic interventions is missing. Thus, the study aims at evaluating the sequential immune status of glioma bearing patients (Astrocytoma Grade I to Grade III) receiving conventional therapeutic measures. The results were compared with the immune status of metastatic secondary glioma and meningioma patients where there is minimal immune suppression and the effect of therapeutic intervention on the above score. METHODS: Functional immune parameters of peripheral blood lymphocytes were assayed by CD2 receptors enumeration through E-rosetting and lymphocyte cytotoxicity assay and assessing the generation of reactive oxygen species by NBT assay of peripheral blood macrophages in patient groups bearing Astrocytoma (Grade I to Grade III), meningioma and secondary glioma. RESULTS: Patients bearing Astrocytoma (all 3 grades) showed maximum immune suppression as compared to the normal subjects, diseased meningioma controls, and secondary glioma. Therapeutic interventions viz. radiotherapy, surgery and radiotherapy after surgery and chemotherapy could not recover the suppressed activity of the CD2 bearing lymphocytes and that of peripheral blood macrophages. However, therapeutic scheduling could recover the functional activity of the CD8 bearing lymphocytes and the CD56 NK cells from that of tumor bearing patients. CONCLUSION: Astrocytoma and not meningioma is capable of causing immunesuppression. As the tumor progresses from Grade I to Grade III, a linear reduction in the functional efficacy of immunocytes is seen to occur. Radiotherapy, surgery, and chemotherapy also induces an inhibitory effect towards the host immune system. The inhibitory effect of tumor as well as of therapy were mainly directed towards the CD2 bearing lymphocyte population and the peripheral blood macrophage population.

Antineoplastic Agents↗