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Dimerization of the RamC Morphogenetic Protein of Streptomyces coelicolor. Michael E. Hudson, 2004.RamC is required for the formation of spore-forming cells called aerial hyphae by the bacterium Streptomyces coelicolor . This protein is membrane associated and has an amino-terminal protein kinase-like domain, but little is known about its mechanismof action . In this study we found that the presence of multiplecopies of a defective allele of ramC inhibits morphogenesisin S . coelicolor, consistent with either titration of a targetor formation of inactive RamC multimers . We identified a domainin RamC that is C terminal to the putative kinase domain andforms a dimer with a Kd of ~0.1 µM . These data suggestthat RamC acts as a dimer in vivo. Role of Positional Hydrophobicity in the Leishmanicidal Activity of Magainin 2. Esther Guerrero, 2004.The emergence of membrane-active antimicrobial peptides as new alternatives against pathogens with multiantibiotic resistance requires the design of better analogues . Among the different physicochemical parameters involved in the optimization of linear antimicrobial peptides, positional hydrophobicity has recently been incorporated . This takes into consideration the concept of the topological distribution of hydrophobic residues throughout the sequence rather than the classical concept of hydrophobicity as a global parameter of the peptide, calculated as the summation of the individual hydrophobicities of the residues . In order to assess the contribution of this parameter to the leishmanicidal mechanisms of magainin 2 analogues, the activities of two of these analogues, MG-H1 (GIKKFLHIIWKFIKAFVGEIMNS) and MG-H2 (IIKKFLHSIWKFGKAFVGEIMNI), which have similar charges, amino acid compositions, and hydrophobicities but different positional hydrophobicities, against Leishmania donovani promastigotes were assayed (T . Tachi, R . F . Epand, R . M . Epand, and K . Matsuzaki, Biochemistry 41:10723-10731, 2002) . The activities were compared with that of the parental peptide, F5W-magainin 2 (GIGKWLHSAKKFGKAFVGEIMNS) . The three peptides were active at micromolar concentrations, in the order MG-H2 > MG-H1 > F5W-magainin 2 . These activities differ from their hemolytic and bactericidal activities . The results demonstrate that positional hydrophobicity, which reflects the presence of short stretches of sequences rich in hydrophobic amino acids, plays an important role in the activities of leishmanicidal peptides . Dam- and OxyR-Dependent Phase Variation of agn43: Essential Elements and Evidence for a New Role of DNA Methylation. Anu Wallecha, 2002.Phase variation of the outer membrane protein Ag43 in E . coli requires deoxyadenosine methylase (Dam) and OxyR . Previously, it was shown that OxyR is required for repression of the Ag43-encoding gene, agn43, and that Dam-dependent methylation of three GATC target sequences in the regulatory region abrogates OxyR binding . Here we report further characterization of agn43 transcription and its regulation . Transcription was initiated from a  70-dependent promoter at the G residue of the upstream GATC sequence . Template DNA and RNA polymerase were sufficient to obtain transcription in vitro, but DNA methylation enhanced the level of transcription . Analyses of transcription in vivo of agn'-lacZ with mutated Dam target sequences support this conclusion . Since methylation also abrogates OxyR binding, this indicates that methylation plays a dual role in facilitating agn43 transcription . In vitro transcription from an unmethylated template was repressed by OxyR(C199S), which resembles the reduced form of OxyR . Consistent with this and the role of Dam in OxyR binding, OxyR(C199S) protected from DNase I digestion the agn43 regulatory region from -16 to +42, which includes the three GATC sequences . Deletion analyses of the regulatory region showed that a 101-nucleotide region of the agn43 regulatory region containing the promoter and this OxyR binding region was sufficient for Dam- and OxyR-dependent phase variation
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