This article traces back thirty years of biotechnology research sponsored by the European Union (EU). It outlines the crucial role played by De Nettancourt, Goffeau and Van Hoeck to promote and prepare the first European programme on biotechnology (1982-1986) run by the European Commission. Following this first biotechnology programme, others followed until the current one, part of the seventh Framework Programme for Research, Technological Development and Demonstration (2007-2013) (FP7). Particular attention is given to the statutory role of the European institutions in the design and orientation of the successive biotechnology programmes, compared to the more informal-yet visionary-role of key individuals upstream to any legislative decision. Examples of success stories and of the role of the biotechnology programmes in addressing societal issues and industrial competitiveness are also presented. Finally, an outline of Horizon 2020, the successor of FP7, is described, together with the role of biotechnology in building the bioeconomy.

The single chain variable fragment (scFv) of an anti-idiotypic catalytic monoclonal antibody, 9G4H9, displaying a β-lactamase-like activity was cloned. The recombinant protein was expressed through the periplasm in Escherichia coli in the presence or in the absence of FkpA, a chaperone-like enzyme and tested for its hydrolytic activity. The results show that the catalytic parameters for hydrolysis of ampicillin by scFv9G4H9 are clearly influenced by the presence of FkpA, indicating that the correct folding of the fragment represents a crucial step for catalysis.

We have induced a polyclonal IgG that degrades the HIV-1 surface antigen, glycoprotein gp120, by taking advantage of the susceptibility of SJL mice to a peptide-induced autoimmune disorder, experimental autoimmune encephalomyelitis (EAE). Specific pathogen-free SJL mice were immunized with structural fragments of gp120, fused in-frame with encephalitogenic peptide MBP(85-101). It has resulted in a pronounced disease-associated immune response against antigens. A dramatic increase of gp120 degradation level by purified polyclonal IgG from immunized versus nonimmunized mice has been demonstrated by a newly developed fluorescence-based assay. This activity was inhibited by anti-mouse immunoglobulin antibodies as well as by Ser- and His-reactive covalent inhibitors. A dominant proteolysis site in recombinant gp120 incubated with purified polyclonal IgG from immunized mice was shown by SDS-PAGE. The SELDI-based mass spectrometry revealed that these antibodies exhibited significant specificity toward the Pro484-Leu485 peptide bond. The sequence surrounding this site is present in nearly half of the HIV-I variants. This novel strategy can be generalized for creating a catalytic vaccine against viral pathogens.

Caging of bovine haemoglobin with increasing amounts of 1-(2-nitrophenyl)ethyl (NPE) and uncaging after a 366 nm irradiation was examined. Caged and photolysed conjugates were characterised by enzymatic assay of the ABTS oxidation, UV/Vis absorbance, and electrospray mass ionisation. Modification of haemoglobin with 50, 75, and 100 equivalents of 1-(2-nitrophenyl)diazoethane led to a progressive decrease of enzymatic activity. Photolysis at 366 nm during 5, 15, and 30 min induced the recovery of a part of the enzymatic activity. ESI analyses showed that a reversible binding of up to 6 NPE groups per alpha-chain and that the removal of most of the photolabile groups occurred rapidly after 5 min of illumination at 366 nm and reached near completion after 15 min. A variable alteration of haemoglobin after labelling could explain that the complete removal of NPE groups did not restore its full oxidative activity.

Mycobacterium sp. strain RP1 degrades morpholine, piperidine, and pyrrolidine and is able to use these compounds as the sole source of carbon, nitrogen, and energy. Cytochrome P450 (MorA) is involved in the biodegradation of these secondary amines. A 3.9-PstI genomic DNA fragment, containing the gene encoding MorA, was cloned and sequenced. Four open reading frames were detected on this DNA fragment. The first encoded a cytochrome P450 designated as MorA which was the second member of the CYP151 family and was named CYP151A2. The second open reading frame (morB) featured a [3Fe-4S] type of ferredoxin. A third gene (morC), exhibiting sequence identity to known reductases, and a fourth truncated gene encoding a putative glutamine reductase (orf1' ), were found downstream of morB. Recombinant MorA cytochrome P450 was purified to homogeneity from Escherichia coli. The purified enzyme was a monomeric soluble protein with an apparent Mr of about 45,000. CYP151A2 catalyzed the ring cleavage of the secondary amines and the Vmax/KMapp values indicated that pyrrolidine is the preferred substrate for this monooxygenase.

An abzyme (9G4H9) displaying a beta-lactamase activity was obtained through the idiotypic pathway and has been previously described. Analysis of the catalytic mechanism by kinetic measurements with various substrates, chemical modifications, and three dimensional modeling, led us to conclude that IgG 9G4H9 displays an activity at the crossroads of beta-lactamases and penicillin-binding proteins (PBPs). Penicillin-binding proteins and beta-lactamases are two closely related enzymes arising from a common ancestral gene. We herein propose that the idiotypic network has allowed the generation of a catalyst with unique catalytic properties, but that could behave as an intermediate between both enzymes.

A library of random peptide sequences was used to select peptides that inhibit an anti-idiotypic catalytic Ig, immunoglobulin (IgG) 9G4H9, with a beta-lactamase-like activity. This library displays cyclic heptapeptides on the surface of bacteriophages and represents a collection of up to 4.5 x 10(9) peptides. The first selection step aimed at enriching the library in species that bind to the whole Ig molecule. The second step was to discriminate peptides that bind to part of the molecule other than the active site. Selected peptides were then screened by surface plasmon resonance analysis. Those displaying measurable K (d) values were assayed for their ability to inhibit the catalytic Ig.

The catalytic mechanism of an anti-idiotypic antibody, 9G4H9, displaying a beta-lactamase activity was investigated. Kinetics experiments suggest that some penicillinic derivatives behave both as substrates and inactivators. Biochemical and immunological experiments strongly indicate that ampicillin may be regarded as a suicide substrate for hydrolysis by 9G4H9. The anti-idiotypic network appears as a way to create enzyme mimics with modified catalytic activities.

A monoclonal antibody 9G4H9 that exhibits a beta-lactamase-like activity was previously obtained in accordance with the idiotypic network theory. This abzyme presents the most catalytic efficiency in amidase activity described in literature (kcat = 0.9 min-1). Some reports have demonstrated that functionality as complex as catalysis may be mimicked in this way. Comparison of the catalytic properties of both enzyme and abzyme previously allowed us to obtain better knowledge about 9G4H9 abzymatic machinery. In attempt to characterize this abzyme, the variable regions of kappa and heavy chain were cloned. We present a 'universal' method to clone the correct Vkappa gene to bypass aberrant Vkappa (abVkappa) produced by MOPC-21-derived hybridomas. Sequences obtained are compared in the GenBank database. The VH and Vkappa genes present some important sequence homology with autoantibodies suggesting a direct relationship between catalytic anti-idiotypic antibody and autoimmunity.

The physiological behaviour of Pseudomonas fluorescens strain R2fN was compared to that of transconjugants [R2fN(RP4)], and two aggregation phenotypes were identified (Agr− and Agr+). Agr+ phenotype is characterized by the appearance of macroscopic aggregates when cells are growing in liquid media. Transconjugants exhibited Agr+ phenotype whereas wild type strain represented Agr−. Evidence is presented to support correlation between Agr+ phenotype acquisition and the presence of the broad-host range plasmid RP4 in strain R2fN. In addition, according to bacterial adherence to hydrocarbon test the transconjugant cells appeared to be very hydrophilic whereas wild type R2fN cells were hydrophobic.

A catalytic IgG (Ab2) displaying a beta-lactamase-like activity was previously obtained by using the antiidiotypic pathway: the particularity of this antibody is that it is a true antiidiotype of the beta-lactamase active site. We have previously demonstrated that this IgG has retained some of the structural information displayed by the beta-lactamase active site, evident from data that polyclonal anti-Ab2 antibodies (Ab3) recognize beta-lactamase. In this article, we investigated the catalytic mechanisms of the abzyme compared to that of the enzyme. The experimental data, allow us to draw hypothesize the catalytic residues required for catalysis.

Transformed callus cultures of Nicotiana tabacum were generated in which the SAM-1 gene from Arabidopsis thaliana encoding S-adenosylmethionine synthetase (SAM-S), under the control of the 35S promoter, had been integrated. The presence of the SAM-1 gene was detected in all tested transformants and the SAM-S activity correlated with the accumulation of SAM in the tobacco callus cultures. Three distinct phenotypic classes were identified among the transgenic cell lines in relation to growth of the cells, structure of the calli, and level of SAM. Transgene silencing was observed in several cultivated transgenic calli and this phenomenon was correlated directly with a low level of SAM-1 mRNA accompanied by a decrease of the SAM-S activity. The transgenic calli overexpressing the SAM-1 gene accumulated a high SAM level. The modifications in SAM-S activity were reflected in the pattern of secondary products present in the different cell lines, thereby demonstrating that the flux through the biosynthetic pathway of a plant secondary product can be modified by means of genetic engineering.

The effect of glucose on the α-amylase production by Bacillus subtilis ATCC-21556 was studied. Initial glucose concentrations up to 20 g/L were found to be directly proportional to the specific α-amylase production in an immobilized-cell batch system, whereas a free-cell batch system presented an inversely proportional relationship with the initial glucose concentration. This might be owing to the α-amylase repression by the glucose present in the culture medium. Three hundred eighty-five percent of the specific α-amylase production with the free-cell system was produced by the immobilized-cell batch culture.

Research Articles| July 22 2010 Prolegomenon Subject Area: Cardiovascular System , Further Areas J.T. Shepherd J.T. Shepherd Search for other works by this author on: This Site PubMed Google Scholar Blood Vessels (1990) 27 (2-5): 70–72. https://doi.org/10.1159/000158798 Article history Published Online: July 22 2010 Content Tools Views Icon Views Article contents Figures & tables Video Audio Supplementary Data Peer Review Share Icon Share Facebook Twitter LinkedIn Email Tools Icon Tools Get Permissions Cite Icon Cite Search Site Citation J.T. Shepherd; Prolegomenon. Blood Vessels 31 December 1990; 27 (2-5): 70–72. https://doi.org/10.1159/000158798 Download citation file: Ris (Zotero) Reference Manager EasyBib Bookends Mendeley Papers EndNote RefWorks BibTex toolbar search Search Dropdown Menu toolbar search search input Search input auto suggest filter your search All ContentAll JournalsJournal of Vascular Research Search Advanced Search This content is only available via PDF. 1990Copyright / Drug Dosage / DisclaimerCopyright: All rights reserved. No part of this publication may be translated into other languages, reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying, recording, microcopying, or by any information storage and retrieval system, without permission in writing from the publisher.Drug Dosage: The authors and the publisher have exerted every effort to ensure that drug selection and dosage set forth in this text are in accord with current recommendations and practice at the time of publication. However, in view of ongoing research, changes in government regulations, and the constant flow of information relating to drug therapy and drug reactions, the reader is urged to check the package insert for each drug for any changes in indications and dosage and for added warnings and precautions. This is particularly important when the recommended agent is a new and/or infrequently employed drug.Disclaimer: The statements, opinions and data contained in this publication are solely those of the individual authors and contributors and not of the publishers and the editor(s). The appearance of advertisements or/and product references in the publication is not a warranty, endorsement, or approval of the products or services advertised or of their effectiveness, quality or safety. The publisher and the editor(s) disclaim responsibility for any injury to persons or property resulting from any ideas, methods, instructions or products referred to in the content or advertisements. Article PDF first page preview Close Modal You do not currently have access to this content.

Because microorganisms frequently live in an immobilized state in natural habitats, a cell-confined system was used to study bacterial conjugation. Two Pseudomonas putida strains were introduced together within calcium alginate gels. Different alginate beads were designed by varying the polysaccharide and the gelation solution concentrations. Microscopic examinations showed that 2% gels were quite homogeneous, but that 1.5% and 1% gels were rather heterogeneous. In these two last cases, shaft-shaped macrostructures were present. They were colonized during the culture by great densities of highly motile bacteria. Gene transfers due to conjugation were investigated in such alginate gel bead microcosms, in batch and continuous cultures. High-initial transfer frequencies were detected whatever the gel, but no conjugation events seemed to occur with further growth in the beads. Transfer frequency values were roughly similar in the different tested systems. Alginate gels used as artificial microcosms may be valuable to study the effect of cell microenvironment on genetic transfers in complex systems.

Catalytic antibodies (abzymes) were developed as tools in chemistry or studied in relation to pathology, but little attention has been paid to their possible biological role.

Antigen mimicry by anti‐idiotypic antibodies is investigated as a reliable strategy to achieve molecular imprinting of an enzymatic activity. A monoclonal anti‐idiotypic antibody (Ab2‐9G4H9) was elicited by using a monoclonal antibody (Ab1‐7AF9) specific for the β‐lactamase active site. Catalytic features of Ab2 were characterized with β‐lactamase substrates. The antibody combining site appeared to have retained a part of the catalytic specificity. The relevance of the idiotypic mimicry concept for the generation of catalytic antibodies was further demonstrated by eliciting a third generation antibody (Ab3), which was shown to recognize β‐lactamase: the complete internal image properties of Ab2 9G4H9, including binding and catalytic properties, were thus checked.—Avalle, B., Thomas, D., Friboulet, A., Functional mimicry: elicitation of a monoclonal antiidiotypic antibody hydrolizing β‐lactams. FASEB J. 12, 1055–1060 (1998)