Biomedical subjects

M S Payne

Publications and source records attributed to M S Payne.

3 recordsLinked to original sources

PubMed1991

Use of alkaline phosphatase fusions to study protein secretion in Bacillus subtilis.

We have constructed a vector designed to facilitate the study of protein secretion in Bacillus subtilis. This vector is based on a translational fusion between the expression elements and signal sequence of Bacillus amyloliquefaciens alkaline protease and the mature coding sequence for Escherichia coli alkaline phosphatase (phoA). We show that export of alkaline phosphatase from B. subtilis depends on a functional signal sequence and that alkaline phosphatase activity depends upon secretion. The vector design facilitates the insertion of heterologous coding sequences between the signal and phoA to generate three-part translational fusions. Such phoA fusions are easily analyzed by monitoring alkaline phosphatase activity on agar plates or in culture supernatants or by immunological detection. Exploitation of this methodology, which has proven to be extremely useful in the study of protein secretion in E. coli, has a variety of applications for studying protein secretion in B. subtilis.

Alkaline Phosphatase↗

PubMed1987

Complement activation by hydroxyethylmethacrylate-ethylmethacrylate copolymers.

Certain biomaterials, including nylon oxygenator and cellulosic dialysis membranes, are potent activators of human complement. In this study, the effect of polymers containing 2-hydroxyethyl methacrylate (HEMA) on the human complement system was investigated. Copolymers of HEMA with ethylmethacrylate (EMA) varying from 100 to 40% HEMA in the monomer made by radiation initiation were used to coat glass discs. These were equilibrated with human plasma in vitro and the degree of complement activation was quantitated by C3a radio immunoassay. Significant activation was caused by copolymers made from monomers containing 60% or greater HEMA. A direct relationship between the amount of activation and the percentage of HEMA was found. The degree of activation by poly HEMA, when corrected for surface area, was quite similar to that observed for dialysis and oxygenator membranes. Similar observations were made when solid casts of crosslinked HEMA/N-vinylpyrrolidone (NVP) copolymer gels were tested, but the magnitude of activation was much greater. The results are significant because complement activation may play an important role in the response to foreign surfaces, in both extravascular and intravascular settings. A new concept of molecular biocompatibility is proposed in which surfaces eliciting molecular transformations in any of the biological defense systems are deemed nonbiocompatible. By this criterion, the hydrogel poly-HEMA, which has so frequently been thought of as biocompatible, is considered to be a molecularly non-biocompatible material.

Biocompatible Materials↗

PubMed1987

Equal stabilities of normal beta globin and nontranslatable beta0 -39 thalassemic transcripts in cell-free extracts.

Patients with beta zero thalassemia arising from premature terminator codon mutations in the gene for beta globin do not produce beta globin protein; these individuals also exhibit a decreased amount of beta globin mRNA in their erythroid cells. The absence of beta globin protein is readily explained by the inability of the beta zero-39 mRNA to be translated. The decrease in beta globin mRNA has been attributed to either decreased cytoplasmic stability of the nontranslatable decreased cytoplasmic stability of the nontranslatable mRNA or to an undefined nuclear lesion. To compare directly the relative stabilities of normal and beta zero-39 thalassemic globin transcripts, we prepared normal and thalassemic beta globin pre-mRNAs and mRNAs using cloned DNA templates and the SP6 promoter-polymerase system. The stability of the transcripts was assessed by incubation in various cell-free extracts. Our results indicate that although the stabilities of the beta globin transcripts varied considerably from one extract to another the stabilities of the beta zero-39 thalassemic pre-mRNAs and mRNAs were equal to those of normal beta globin mRNAs in every extract tested.

Animals↗