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Abstract: Some scorpion venom contain heterodimeric phospholipases A2. They were shown to be toxic to insects and to cause edema and/or hemolysis of mammalian erythrocytes. This manuscript describes the results of cDNA cloning of five different heterodimeric phospholipases from the venomous glands of the Mexican scorpion Anuroctonus phaiodactylus. The amino acid sequence deduced from the heterodimeric phospholipases open reading frames corresponds in each case to a different isoform. The nucleotide sequences corresponding to two of these genes were also obtained by directly sequencing genomic DNA. The cDNA isoforms show high similarity with the heterodimeric phospholipase Phaiodactylipin purified from the same scorpion. However, similar phospholipases were also found in scorpions from other species and the sequences available were used to construct a phylogenetic tree. In order to understand better the gene structure and phylogeny of these enzymes we analyzed their sequences and compared them with secretory phospholipases of other sources from groups I, II and III. The genomic DNA sequence of a similar phospholipase from bee venomous glands was also cloned. The information available on a Drosophila phospholipase was included in this analysis. The phospholipases of groups I and II contain a conserved exon–intron structure (four or five exons of the mature segment of the enzyme are separated by three or four introns). Also, the gene structure of the phospholipases from A. phaiodactylus and that of the bee venom, belonging to group III phospholipases, are interrupted by three introns. The mature peptide of the bee enzyme is a single polypeptide chain, coded by four exons, whereas those from the scorpion studied here although having four exons, showed the presence of two different polypeptides in its native state. The mature protein is processed after synthesis, producing the heterodimeric structure: a long and a short-peptide chain, linked by a disulfide bridge. The small subunit is the one coded by the fourth exon. The human phospholipase A2 and that of Drosophila, also classified into the group III phospholipases, have a considerably different exon–intron organization. [Copyright &y& Elsevier]

Summary: Partial nucleotide sequences of the citrate synthase (gltA) gene from different rhizobia genera were determined. Tree topologies based on this housekeeping gene were similar to that obtained using 16S rRNA sequences. However gltA appeared to be more reliable at determining phylogenetic relationships of closely related taxa. We propose gltA sequences as an additional tool to be used in molecular phylogenetic studies. [Copyright &y& Elsevier]

Abstract: Venom components from the centipede Scolopendra viridis Say were studied, using both the soluble venom and a cDNA library prepared from mRNA of the venomous glands. Separation of the soluble venom by high performance liquid chromatography (HPLC) permitted to obtain at least 54 different fractions. The fraction eluting at 46.24min showed phospholipase activity. The enzyme was purified to homogeneity and the first 25 amino acid residues were identified by Edman degradation. From the cDNA library several genes were cloned, one of which codes for a protein with identical amino acid sequence as the one experimentally determined. The cloned gene codes for a signal peptide of 28 amino acids and a mature peptide of 119 residues. The molecular weight of the enzyme was estimated by mass spectrometry and shown to be 13,752Da, which matches exactly with the molecular mass expected from the deduced amino acid sequence of the gene. Phylogenetic analysis of this sequence, in comparison with other known from venomous animals, showed that it is more similar to snake phospholipases than to insect or arachnid sequences, suggesting that it has been submitted to convergent evolution. To the best of our knowledge this is the first time that a phospholipase from this species of animal is fully characterized. We have named it Scol/Pla. [Copyright &y& Elsevier]

Abstract: The resistance-nodulation-cell division (RND) protein family is a ubiquitous group of proteins primarily present in bacteria. These proteins, involved in the transport of multiple drugs across the cell envelope in bacteria, exhibit broad substrate specificity and act like efflux pumps. In this work, a protein belonging to the RND protein family, AcrB of Escherichia coli was used as a working model to predict in silico the compounds transported by 47 RND proteins. From AcrB we extracted and clustered 14 amino acids directly involved in substrate interactions. This clustering provides enough information to identify 16 groups that correlates with the ligand they extrude, such as proteins expelling aromatic hydrocarbons (SrpB cluster) or proteins expelling heavy metals (CnrA cluster). The relationship between conserved, cluster-specific and variable residues indicates that although the ligand-binding domain is conserved in structure, it has enough flexibility to recognize specifically a diversity of molecules. [Copyright &y& Elsevier]

Abstract: A Leuconostoc mesenteroides NRRL B-512 F levansucrase gene, (levS), was isolated, sequenced and cloned in Escherichia coli. The recombinant enzyme was shown to be a fructosyltransferase producing a polymer identified by 13C-NMR as levan. Based on sequence analysis, we found that this levansucrase is a mosaic protein, bearing structural features of glucosyltransferases in the amino and carboxy terminal regions similarly to inulosucrase from Leuconostoc citreum. The phylogenetic analysis of the C-terminal region domain of levansucrases from L. mesenteroides demonstrates that they group together into a novel putative sub-family of genes and evolved long before all other glucosyltransferases, while their catalytic domain structure is species related. [Copyright &y& Elsevier]

Abstract: We have addressed the distribution and abundance of 75 transcription factor (TF) families in complete genomes from 90 different bacterial and archaeal species. We found that the proportion of TFs increases with genome size. The deficit of TFs in some genomes might be compensated by the presence of proteins organizing and compacting DNA, such as histone-like proteins. Nine families are represented in all the bacteria and archaea we analyzed, whereas 17 families are specific to bacteria, providing evidence for regulon specialization at an early stage of evolution between the bacterial and archeal lineages. Ten of the 17 families identified in bacteria belong exclusively to the proteobacteria defining a specific signature for this taxonomical group. In bacteria, 10 families are lost mostly in intracellular pathogens and endosymbionts, while 9 families seem to have been horizontally transferred to archaea. The winged helix-turn-helix (HTH) is by far the most abundant structure (motif) in prokaryotes, and might have been the earliest HTH motif to appear as shown by its distribution and abundance in both bacterial and archaeal cellular domains. Horizontal gene transfer and lineage-specific gene losses suggest a progressive elimination of TFs in the course of archaeal and bacterial evolution. This analysis provides a framework for discussing the selective forces directing the evolution of the transcriptional machinery in prokaryotes. [Copyright &y& Elsevier]