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J Marzillier

Publications and source records attributed to J Marzillier.

8 recordsLinked to original sources
PubMed1994

Efficient production of active and mutated ADP-ribosyltransferase (S1) of pertussis toxin using affinity expression cassette polymerase chain reaction.

We describe an efficient, general approach for cloning, expression and purification of heterologous proteins in Escherichia coli host strains. The affinity expression cassette polymerase chain reaction (AEC-PCR) allows the insertion of virtually any coding sequence in suitable expression vectors. For ease of purification of the (over)produced protein the gene expression cassettes are engineered by specifically designed oligonucleotide primers in the polymerase chain reaction (PCR) to contain either 3' or 5' additional nucleotides coding for a short amino acid sequence constituting an 'affinity block' fused to either end of the protein. This allows a one-step purification by affinity chromatography. In combination with PCR-mediated site-specific mutagenesis this approach is a highly efficient way for the expression and isolation of proteins and for the analysis and dissection of functional domains. The application of AEC-PCR is demonstrated by the cloning, production and purification of the native, active and the mutagenized, inactive ADP-ribosyltransferase (S1 subunit) of pertussis toxin. In this example, a string of six histidines has been engineered to either the N-terminal or the C-terminal end of the protein to serve as 'affinity block' for the isolation of the recombinant protein by immobilized metal ion affinity chromatography (IMAC). Thus, the S1 subunit can now be produced in sufficient quantities to facilitate further studies on its immunological and molecular properties.

Base Sequence↗
PubMed1991

Identification of linear B-cell determinants of pertussis toxin associated with the receptor recognition site of the S3 subunit.

Receptor recognition of pertussis toxin is mediated by the B oligomer consisting of subunits S2, S3, 2xS4, and S5. One possible way to interfere with toxin action would be the inhibition of recognition and binding of the cellular receptor(s) by preformed toxin-directed antipeptide antibodies. A prerequisite for this approach is the localization of linear antigenic determinants followed by the identification of inhibitory epitopes. Anti-S2 peptide antibodies have been shown to inhibit binding of the holotoxin to in vitro model receptor systems. For the elucidation of linear antigenic and immunogenic determinants harbored in the S3 subunit, synthetic peptides corresponding to selected linear amino acid sequences of S3 have been prepared and used to raise peptide-specific antibodies in rabbits. All peptides elicited a strong homologous response. Four synthetic peptides reacting with anti-pertussis toxin antibodies (R36-51, R87-95, R134-150, and R147-160) have been identified. Seven synthetic peptides (R1-12, R12-23, R14-29m, R36-51, R95-107, R134-150, and R164-178) induced antibodies recognizing pertussis toxin. Thus, these segments correspond to linear antigenic determinants. Analogous to the S2 subunit, the N terminus of S3 proved to be immunorecessive in the native toxin. The highly homologous S2 subunit was only bound strongly in Western blotting (immunoblotting) by antiserum directed at peptide R164-178, which is identical in the S2 and S3 subunits. A weak recognition of S2 in Western blotting was observed with anti-R95-107 antiserum. The ability of affinity-purified anti-S3 peptide antibodies to interfere with pertussis toxin binding was investigated by hemagglutination of goose erythrocytes as a model receptor system for S3-mediated receptor recognition. Antipeptide antibodies directed at R1-12, R12-23, R14-29m, and R36-51 inhibited hemagglutination of goose erythrocytes. This indicates that the corresponding antigenic regions in the S3 subunit are associated with the formation of the receptor binding domain. Inhibition of B-oligomer-mediated pertussis toxin binding to cellular receptors by preformed antipeptide antibodies of sufficient affinity should not only block the detrimental effects of the S1 subunits, but also interfere with the mitogenic effects attributed to the B oligomer.

Amino Acid Sequence↗
PubMed1990

Development and evaluation of a training course for practice nurses: helping people to stop smoking.

This paper describes a 2-day training course for practice nurses in advice-giving to smokers. The course incorporated information on the epidemiology, physiology and psychology of smoking and a strategy for giving advice based upon the health-beliefs model. The course was systematically evaluated on measures of knowledge about health and smoking and on confidence in various advice-giving activities. The results showed that the nurses gained significantly in both knowledge and confidence following the course. The prospect of further training courses for practice nurses in other areas of health promotion is discussed.

Attitude of Health Personnel↗
PubMed1989

Randomized controlled trial of anti-smoking advice by nurses in general practice.

Practice nurses are playing an increasingly prominent role in preventive care, including the provision of anti-smoking advice during routine health checks. A randomized controlled trial was designed to assess the effectiveness of anti-smoking advice provided by nurses in helping smokers to stop smoking. A total of 14,830 patients aged 16-65 years from 11 general practices completed a brief questionnaire on general health, including smoking status, at surgery attendance. The doctor identified 4330 smokers and randomly allocated 4210 to control or intervention groups. The doctor asked those in the intervention group to make an appointment with the practice nurse for a health check. The attendance rate at the health check was 26%. Smokers were sent follow-up questionnaires at one month and one year, and those who did not respond to two reminders were assumed to have continued to smoke. There was no significant difference in reported cessation between the intervention and control groups at one month or one year. However, there was a significant difference in the proportion of patients who reported giving up within one month and who had not lapsed by one year--0.9% in controls and 3.6% in the intervention group (P less than 0.01). Nevertheless, the effect of the nurse intervention itself may be small as the sustained cessation rate in attenders was only 42.4% higher than in non-attenders. The deception rate in reporting cessation, as measured by urinary cotinine, was of the order of 25%.

Adolescent↗
PubMed1988

Smoke screens.

Explore the source record for details and available documents.

Community Health Nursing↗
PubMed1988

[Streptozocin-sensitized lymphocytes suppress the glucose-stimulated insulin secretion of the islands of Langerhans].

Isolated islets of Langerhans (from C57Bl/6J mice) were either incubated with culture medium only or medium substituted by 1 mM streptozocin (SZ) and cultivated with lymphocytes (Ly) for seven days. Ly were isolated from the same mouse strain sensitized against SZ by 5 x 40 mg SZ per kg and day intraperitoneally or 1 mg SZ subcutaneously in vivo. Glucose-stimulated insulin secretion rate was used as an indicator for the metabolic condition of islets. SZ-pretreated islets showed significantly decreased insulin secretion in the presence of Ly. The secretion capacity of untreated islets was not impaired.

Animals↗
PubMed1986

Practice nurses and antismoking education.

A questionnaire on antismoking activities and education was sent to 369 nurses in general practice. The response rate was 80%. Although most of the nurses sometimes advised patients about smoking, routine antismoking education occurred less frequently. Only a few regularly referred smokers to other agencies for help, recommended aids to stop smoking, or used antismoking literature. Although the nurses thought that they had an important role in helping smokers to give up, they expressed little confidence in their effectiveness, believing that advice from the general practitioner and the smoker's personal determination to give up have more impact. The nurses expressed a need for training in antismoking education. Seventy seven per cent were interested in attending seminars and listed information about smoking, techniques for stopping, and counselling skills as priorities. If practice nurses are to use opportunities in primary care to help smokers there is clearly a need to provide further training and to establish the effectiveness of nurses in their role as smoking educators.

Attitude of Health Personnel↗