Biomedical subjects

R Weilbaecher

Publications and source records attributed to R Weilbaecher.

3 recordsLinked to original sources

PubMed1998

Yeast transcript elongation factor (TFIIS), structure and function. II: RNA polymerase binding, transcript cleavage, and read-through.

The transcriptionally active fragment of the yeast RNA polymerase II transcription elongation factor, TFIIS, comprises a three-helix bundle and a zinc ribbon motif joined by a linker region. We have probed the function of this fragment of TFIIS using structure-guided mutagenesis. The helix bundle domain binds RNA polymerase II with the same affinity as does the full-length TFIIS, and this interaction is mediated by a basic patch on the outer face of the third helix. TFIIS mutants that were unable to bind RNA polymerase II were inactive for transcription activity, confirming the central role of polymerase binding in the TFIIS mechanism of action. The linker and zinc ribbon regions play roles in promoting cleavage of the nascent transcript and read-through past the block to elongation. Mutation of three aromatic residues in the zinc ribbon domain (Phe269, Phe296, and Phe308) impaired both transcript cleavage and read-through. Mutations introduced in the linker region between residues 240 and 245 and between 250 and 255 also severely impaired both transcript cleavage and read-through activities. Our analysis suggests that the linker region of TFIIS probably adopts a critical structure in the context of the elongation complex.

Models, Molecular↗

PubMed1994

Termination-altering amino acid substitutions in the beta' subunit of Escherichia coli RNA polymerase identify regions involved in RNA chain elongation.

To identify regions of the largest subunit of RNA polymerase that are potentially involved in transcript elongation and termination, we have characterized amino acid substitutions in the beta' subunit of Escherichia coli RNA polymerase that alter expression of reporter genes preceded by terminators in vivo. Termination-altering substitutions occurred in discrete segments of beta', designated 2, 3a, 3b, 4a, 4b, 4c, and 5, many of which are highly conserved in eukaryotic homologs of beta'. Region 2 substitutions (residues 311-386) are tightly clustered around a short sequence that is similar to a portion of the DNA-binding cleft in E. coli DNA polymerase I. Region 3b (residues 718-798) corresponds to the segment of the largest subunit of RNA polymerase II in which amanitin-resistance substitutions occur. Region 4a substitutions (residues 933-936) occur in a segment thought to contact the transcript 3' end. Region 5 substitutions (residues 1308-1356) are tightly clustered in conserved region H near the carboxyl terminus of beta'. A representative set of mutant RNA polymerases were purified and revealed unexpected variation in percent termination at six different rho-independent terminators. Based on the location and properties of these substitutions, we suggest a hypothesis for the relationship of subunits in the transcription complex.

Amino Acid Sequence↗

PubMed1989

Phase I study of cancer therapy with recombinant interleukin-2 administered by intravenous bolus injection.

Sixty-six patients with disseminated malignancy were treated with recombinant interleukin-2 (IL-2) on a three times a week (M, W, F) IV-bolus injection schedule. Doses ranged from 0.001 to 14.0 x 10(6) units/M2 body surface area. Consecutive groups of 3-5 patients were placed on each dose level and were maintained on that level except for dosage de-escalation for toxicity. Toxicity to all major organ systems were noted with major toxicity including fever and chills, anorexia, fatigue and malaise, arthralgias and arthritis as well as hepatic and renal toxicity. All toxicity reversed within one week of drug cessation. Renal toxicity manifested by azotemia, arthritis and fatigue were the common dose limiting toxicities and the maximally tolerated dose was 12 x 10(6) units/M2. Pharmacokinetic studies indicated a short half-life (T1/2 alpha = 7-23 minutes). At doses over 0.5 x 10(6) units/M2 increases in absolute lymphocytes and eosinophil counts were noted. All T lymphocyte subsets increased. Maximal increases were seen at 4-8 x 10(6) units/M2 with a lesser increase at 10-14 x 10(6) units/M2 dosage level. Circulating NK cells also increased while circulating LAK cells were detected during therapy. Partial responses were noted in 3 patients with melanoma. These lasted 4, 6 and 16 months and involved pulmonary, pulmonary plus mesenteric and retro-orbital plus hepatic metastases respectively in these patients.

Drug Evaluation↗