High efficiency intergeneric conjugal transfer of plasmid DNA from Escherichia coli to methyl DNA-restricting streptomycetes

Many streptomycetes, including S. coelicolor A3(2), possess a potent methyl-specific restriction system which can present an effective barrier to the introduction of heterologous DNA. We have compared the efficiency of intergeneric conjugal transfer of different types of plasmids to S. coelicolor and S. lividans 66 using two E. coli donors: the standard, methylation proficient strain S17-1. and the methylation deficient donor, ET12567(pUB307). We demonstrate that the methylation deficient donor can yield > 10⁴-fold more S. coelicolor exconjugants than the standard donor. In the case of pSET152 derivatives, which integrate into the host chromosome by site-specific recombination, up to 10% of streptomycete spores in the conjugation mixture inherit the plasmid. The conjugation procedure is efficient enough to obtain exconjugants with 'suicide' delivery plasmids and therefore provides a simple route for conducting gene disruptions in methyl DNA-restricting streptomycetes, and possibly other bacteria.     

Computational studies on the backbone-dependent side-chain orientation induced by the (S,S)-CXC motif

Disulfide cyclization is a well-known procedure to impose conformational restriction to peptides undergoing backbone flexibility. Rigid conformations are induced only for small rings with a specific combination of amino acids. In this work, we present a computational search of the backbone and backbone-dependent side-chain orientation of two series of linear and cyclic peptide analogs. The -C[XY]C- scaffold (where X,Y is arginine, aspartic acid or alanine residue) in its open and (S,S) cyclic form was used for the design of the studied analogs. Thirty-six compounds, resulting from the extension with one residue at either the N- or the C-terminus were studied with classical MD. The local backbone conformation and the relative orientation of the X and Y side chains induced by either cyclization and/or the presence of the charged residues are discussed. From the present study it is concluded that cyclization has a great impact on the synplanar orientation of the X and Y side chains in the (S,S)Ac-XCYC-NH2 series of compounds while charge-charge interaction has only a weak synergic effect. On the contrary, the antiplanar orientation is favored in the case of (S,S)Ac-CXCY-NH2.     

Conformational requirements for molecular recognition of acetylcholine receptor main immunogenic region (MIR) analogues by monoclonal anti-MIR antibody: A two-dimensional nuclear magnetic resonance and molecular dynamics approach

The conformational properties of two [D -A⁷⁰, A⁷⁶] and [Aib⁷⁰, A⁷⁶] analogues of the α67–76 Torpedo acetylcholine receptor fragment, with low binding capacity for the anti main immunogenic region (MIR) antibodies, were studied in DMSO by two-dimensional nmr techniques and molecular dynamics simulations. The results were compared to the free and bound conformations of the [A⁷⁶] analogue, which has twice more affinity for the anti-MIR monoclonal antibody 6 (mAb6), than the natural Torpedo sequence. It appeared that a single substitution of the A⁷⁰, at a crucial position, by the D -A⁷⁰ or Aib⁷⁰, could modify completely the conformational behavior of the peptide and reduced its recognition by the anti-MIR antibody. The WNPADY rigid structure at the N-terminal part was essential for antibody recognition. The adjacent more flexible C-terminal sequence (GGIK) gives additional stability to the monoclonal antibody–peptide complex probably due to an adequate orientation of the peptide side chains in the complex, by setting them in close contact with the antibody. © 1993 John Wiley & Sons, Inc.     

Cytogenetic and cell-surface marker studies in two non-Hodgkin's lymphomata of T-cell origin

Summary The chromosomes from 2 cases of lymphoma involving proliferation of abnormal T-lymphocytes are described. The tumours differed histologically and chromosomally, although certain of the structural chromosome changes found are of common occurrence in other, mostly B-cell, lymphomata.     

The adult rat leydig cell aromatase activity is stimulated by a sertoli cell factor [Postor 38]

The adult rat Sertoli cell secretes a factor that stimulates the Leydig cell aromatase activity via protein synthesis, at a step beyond the adenylate cyclase. This factor is of proteic nature, species and tissue specific, different from the Sertoli cell LHRH-like compound, and the germ cells exert a negative control on its synthesis.     

Design, synthesis and catalytic activity of a serine protease synthetic model

Summary The design, synthesis and catalytic properties of a cyclic branched peptide carrier that possesses the catalytic triad residues of the serine proteases is reported. The synthesis of the peptide model was totally completed on solid support using three different orthogonal amino protecting groups. Hydrolytic activity measurements against Suc-Ala-Ala-Ala-pNA substrate showed that it is hydrolysed by the peptide model to a small extent. Despite this small hydrolytic activity, it is the first time, to our knowledge, that hydrolysis of such a substrate is reported by an enzyme model compound. Contrary, this enzyme model peptide showed considerable activity against the Boc-Ala-pNP substrate (k _cat =0.414 min⁻¹ andK _m =0.228 mm). These results suggest that the designed carrier brings in appropriate contact the catalytic triad residues (Ser, His, Asp) resulting in the obtained hydrolytic activity.