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M C Schmidt

Publications and source records attributed to M C Schmidt.

At least 55 records · Page 3Linked to original sources
PubMed1989

Yeast TATA-box transcription factor gene.

The first step in the transcription of most protein-encoding genes in eukaryotes is the binding of a transcription factor to the TATA-box promoter element. This TATA-box transcription factor was purified from extracts of the yeast Saccharomyces cerevisiae by using reconstitution of in vitro transcription reactions as an assay. The activity copurified with a protein whose sodium dodecyl sulfate/polyacrylamide gel mobility is 25 kDa. The sequence of the amino-terminal 21 residues of this protein was determined by sequential Edman degradation. A yeast genomic library was screened with mixed oligonucleotides encoding six residues of the protein sequence. The yeast TATA-box factor gene was cloned, and DNA sequencing revealed a 720-base-pair open reading frame encoding a 27,016-Da protein. The identity of the clone was confirmed by expressing the gene in Escherichia coli and detecting TATA-box factor DNA binding and transcriptional activities in extracts of the recombinant E. coli. The TATA-box factor gene was mapped to chromosome five of S. cerevisiae. RNA blot hybridization and nuclease S1 analysis indicated that the major TATA-box factor mRNA is 1.3 kilobases, including an unusually long 5' untranslated region of 188 +/- 5 nucleotides. Homology searches showed a region of distant similarity to the calcium-binding structures of calpains, a structure that has a conformation similar to the helix-turn-helix motif of DNA binding proteins.

Amino Acid Sequence↗
PubMed1989

Sp1 activates transcription without enhancing DNA-binding activity of the TATA box factor.

We have studied the interactions of the Sp1 and IID transcription factors with a simple RNA polymerase II promoter. The adenovirus E1B core promoter consists essentially of a GC box and a TATA box, binding sites for the Sp1 and IID transcription factors, respectively. The E1B promoter is accurately transcribed in vitro using a mammalian transcription system. Sp1 activates E1B transcription in vitro in reactions using IID factor isolated from either human or yeast cells. In DNase I footprinting studies, Sp1 bound rapidly to its recognition sequence even at 0 degrees C (t1/2 less than 1 min). In contrast, yeast IID bound more slowly (t1/2 approximately 6 min at 25 degrees C) and required thermal energy for stable binding to the TATA box sequence. Dissociation rates were measured by the addition of specific oligonucleotide competitors to preformed DNA-protein complexes. Sp1 dissociates rapidly (t1/2 less than 1 min) at 25 degrees C, while yeast IID dissociates with an estimated t1/2 of 1 h at 25 degrees C. Sp1 and yeast IID bound to the E1B promoter simultaneously but independently. The rates of binding and dissociation of these factors were not significantly affected by the presence of the other factor. Bound Sp1 factor did not alter or enhance the yeast IID footprint. Oligonucleotide challenge of in vitro transcription reactions indicated that Sp1 also did not enhance the binding of the human IID factor to the E1B promoter. Thus the Sp1 factor activates transcription of the E1B gene by a mechanism that does not enhance the DNA-binding activity of the IID factor. Sp1 factor activates E1B transcription by 5- to 10-fold in vitro. Under these in vitro transcription conditions, transcripts due to reinitiation from an individual promoter complex contribute only a small portion of the total yield of E1B transcripts. Thus Sp1 cannot activate transcription by increasing the rate of initiation events per complex. Instead it appears that Sp1 acts by increasing the number of productive transcription complexes formed in vitro.

Adenoviridae↗
PubMed1987

nusA protein of Escherichia coli is an efficient transcription termination factor for certain terminator sites.

We have studied the factors that affect transcription termination in vitro at the tR2 terminator of bacteriophage lambda and at the T1 terminator of the Escherichia coli rrnB operon. Termination efficiency at both of these sites is enhanced by the E. coli nusA protein, giving final efficiencies of termination in vitro comparable to those estimated in vivo. Transcripts terminated in the presence of nusA protein are all released from the RNA polymerase complex, indicating that a complete termination reaction is involved, rather than simply induction of a long pause at the terminator. The termination factor activity of the nusA protein does not depend on the presence of rho protein and is not detectably enhanced by that factor. Thus, the nusA protein appears to play a pleiotropic role in E. coli transcription, serving as an antitermination factor, RNA polymerase subunit and true termination factor for some terminator sites.

Bacterial Proteins↗
PubMed1986

Nonenzymatic radiolabeling of protein by 32P-containing nucleotides.

We report a nonenzymatic reaction which results in the radiolabeling of proteins by 32P-containing nucleoside triphosphates. The labeling reaction does not require any cofactors, but is greatly enhanced by the presence of alcohols. Even under optimal conditions, less than 1% of the protein molecules undergo modification. This nonspecific labeling represents a serious artifact which may become significant in systems involving low levels of specific labeling, such as photoaffinity labeling. Since the reaction is not limited to specific proteins, this may, however, provide a simple and rapid procedure for the preparation of labeled proteins.

Adenosine Triphosphate↗
PubMed1984

Binding of rho factor to Escherichia coli RNA polymerase mediated by nusA protein.

The E. coli transcription termination factor rho binds specifically to purified nusA protein. Since nusA protein binds tightly to RNA polymerase, this provides a way of coupling rho to the elongating RNA polymerase complex through protein-protein interactions. These rho-nusA interactions may play a role in modulating rho action at certain terminators and could also be important in the action of antitermination factors such as lambda N protein.

Adenosine Triphosphatases↗
PubMed1984

Amplification and isolation of Escherichia coli nusA protein and studies of its effects on in vitro RNA chain elongation.

The Escherichia coli nusA gene product is an RNA polymerase binding protein which has been implicated in a variety of cellular and viral termination and antitermination processes. To facilitate large-scale preparation and biochemical studies of the nusA protein, we have cloned the nusA gene into a lambda PL-derived overexpression vector. E. coli strains bearing the resulting plasmid (pMS7) produce large amounts of nusA protein when induced, and the protein is easily purified to homogeneity. Biochemical studies of nusA protein reveal that it inhibits in vitro RNA chain elongation by E. coli RNA polymerase with a variety of templates. Two modes of inhibition are found. Inhibition of elongation with poly[d(A-T] ) template is completely competitive with nucleoside triphosphates and shows an inhibitory constant (Ki) of 3 X 10(-7) M. In contrast, inhibition of elongation with T7 DNA as template is mixed. One component of the inhibition is competitive with nucleoside triphosphate substrates and is reversed at elevated substrate concentrations. A second inhibitory component remains even at saturating substrate concentrations; this sequence-dependent mode of inhibition shows a much lower Ki of 2 X 10(-8) M. The existence of two different modes of inhibition might be explained if two molecules of nusA protein can bind to each RNA polymerase complex. The interaction of nusA protein with elongating RNA polymerase molecules is not processive but appears to be characterized by rapid association and dissociation. Under proper conditions, a sigma-nusA cycle [Greenblatt, J., & Li, J. (1981) Cell (Cambridge, Mass.) 24, 421-428] can be demonstrated in vitro in which each polymerase goes through multiple rounds of transcription involving successive interactions with sigma and the nusA protein.

Bacterial Proteins↗
PubMed1980

Localizing functioning thyroid following total thyroidectomy by use of a combination scanning technique.

A technique is described using 1-131-sodium iodine and Tc-99m-sodium pertechnetate for localizing functioning thyroid tissue in the neck following total thyroidectomy. The procedure consists of obtaining a 1-131 thyroid scan. If functioning thyroid tissue is found, it can be localized by means of skin markers as well as a swallow using Tc-99m-sodium pertechnetate. The procedure is safe, simple, and cost efficient and provides a means of localizing functioning thyroid that may influence subsequent therapy.

Adenocarcinoma↗
PubMedDate not supplied

A TATA box implicated in E1A transcriptional activation of a simple adenovirus 2 promoter.

Adenovirus E1A proteins stimulate transcription by RNA polymerases II and III from many promoters. The detailed mechanism of transcriptional activation (transactivation) by E1A proteins remains unclear, but genetic and biochemical results suggest that E1A products might act to stimulate the activity of cellular transcription factors. In this study, a detailed mutational analysis of the adenovirus E1B promoter was undertaken to define the DNA sequences required for proper basal transcription and E1A transactivation. Two key findings emerged: first the E1B promoter is an unusually simple RNA polymerase II promoter requiring only two sequence elements for proper regulation, the TATA box and a binding site for transcription factor Sp1; and second only mutations in the TATA box interfere with E1A-transactivation, suggesting that E1A mediates its effect on this promoter through the TATA-box transcription factor.

Adenovirus Early Proteins↗
PubMedDate not supplied

Infective endocarditis in a patient with Hodgkin's lymphoma: a case report.

A common sequela of certain malignancies is nonbacterial thrombotic endocarditis (NBTE), a phenomenon in which sterile fibrin/platelet aggregates are deposited onto normal cardiac valves. These verrucae represent a predisposing factor for the initiation of infective endocarditis following a bacteremia. This paper presents a case history which is highly suggestive of infective endocarditis which occurred as a result of multiple odontogenic abscesses in a patient with Hodgkin's lymphoma. The case illustrates the important role that the dentist can play in the management of cancer patients and emphasizes a wholistic concept of medical care in which the dentist is an integral member of the health-care team.

Adult↗
PubMedDate not supplied

The International Normalized Ratio as a measure of anticoagulation: significance for the management of the dental outpatient.

Susceptibility to bleeding in the patient treated with coumarin derivatives poses difficult management issues for the dentist. Classic monitoring for anticoagulation relied on the prothrombin time, but reagent variability issues have caused the emergence of the International Normalized Ratio (INR) as an alternative, and possibly more reliable, laboratory test for the assessment of a patient's anticoagulation status. The literature suggests that an INR range of 1.5 to 2.5 represents the most appropriate level of anticoagulation for the surgical patient. That is, at this INR, some protection is afforded against thrombo-embolic events without significantly sacrificing effective post-operative hemostasis. Since the patient with an INR value within the recommended range is still anticoagulated, he/she is still at risk for post-operative bleeding. Adherence to local hemostatic measures--such as pressure, topical thrombin, alveolar-placed resorbable sponges, and primary wound closure--offers the patient important adjunctive protection against post-operative bleeding.

Ambulatory Care↗