Assuntos
Lepidópteros/genética , Neuropeptídeos/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , DNA Complementar/genética , Feminino , Hormônios Juvenis/antagonistas & inibidores , Hormônios Juvenis/biossíntese , Lepidópteros/metabolismo , Masculino , Manduca/genética , Dados de Sequência Molecular , Sondas de Oligonucleotídeos/genéticaRESUMO
The location and structure of the cos ends of bacteriophage D3, which infects Pseudomonas aeruginosa strain PAO, has been determined using a combination of deletion analysis, transposon mutagenesis, and sequencing directly off the phage DNA. Phage D3 was found to have 9-bp 3' cos ends, making it the first phage of a Gram-negative organism known to have 3' cos ends. A 700-bp region flanking the cos site was necessary for efficient transduction of D3 cosmid derivatives. This region was found to contain incomplete inverted repeat sequences flanking the cos site, along with adenine-rich repeats homologous to coliphage gama Ter binding sites. Possible IHF binding sites were also present.
Assuntos
Fagos de Pseudomonas/genética , Sequência de Bases , Clonagem Molecular , Meios de Cultura , DNA Viral , Dados de Sequência Molecular , Mutagênese Insercional , Pseudomonas aeruginosa/virologia , Mapeamento por Restrição , Deleção de Sequência , Homologia de Sequência do Ácido NucleicoRESUMO
A 15-residue neuropeptide, Manduca sexta allatostatin (Mas-AST), strongly inhibits juvenile hormone (JH) biosynthesis in vitro by corpora allata (CA) from Manduca fifth-stadium larvae and adult females as well as Helicoverpa zea adult females (Kramer et al., 1991 Proc. Natl. Acad. Sci (USA) 88, 9458-9462). In contrast, this study found that 1.0 microM Mas-AST has no JH biosynthesis inhibitory activity in Pseudaletia unipuncta sixth instar larvae or newly-emerged (day 0) adults but inhibited CA of 5-day-old adult females by 60%. From a P. unipuncta brain cDNA library, was isolated a cDNA that encodes a 125 amino acid polypeptide containing the Mas-AST sequence. Within the precursor, Mas-AST is situated at the carboxy terminus and is flanked by different dibasic proteolytic cleavage signals. The Pseudaletia gene specifying the Mas-AST peptide is present as a single copy per haploid genome. Expression of this gene was low in Pseudaletia sixth instar larvae, prepupae and early pupae but was relatively high in late pupae, and day 1 and 3 adults of both sexes. In day 5 adults, the relative transcript level appears to be maintained in females but declines in males. This pattern of Mas-AST expression does not correlate well with the profile of JH biosynthesis in Pseudaletia, which increases during the first 5 days of adult life, suggesting additional or alternative functions for this peptide.
Assuntos
Proteínas de Insetos , Mariposas/genética , Peptídeos/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , DNA Complementar , Feminino , Expressão Gênica , Hormônios Juvenis/antagonistas & inibidores , Hormônios Juvenis/biossíntese , Masculino , Manduca/metabolismo , Dados de Sequência Molecular , Neuropeptídeos/genética , Precursores de Proteínas/genética , Ácido Pirrolidonocarboxílico/análogos & derivados , RNA MensageiroRESUMO
The nucleotide sequence of the replicative origin (OriR) region of the small cryptic broad-host-range plasmid, pRO1600, which forms the basis of a number of useful cloning vectors has been determined. In addition it has been subjected to Tn5 mutagenesis, deletion analysis, and subcloning in order to define the regions essential for replication in Pseudomonas aeruginosa. The sequence (1894 bp) contains a fragment derived from transposon Tn1. The OriR region is structurally related to other replication (Rep) protein-dependent origins in that it has an A-T-rich region upstream of four 17-bp direct repeats (iterons) which presumably function in initiator protein binding. The sequence also contains a DNA-A-binding site and an open reading frame which could encode a basic (pI 10.6) 25,343-Da Rep protein with homology to RepA from the Neisseria gonorrhoeae beta-lactamase plasmid pFA3. The possible evolutionary origin of this plasmid in P. aeruginosa (RP1) is discussed.
Assuntos
Proteínas de Bactérias/genética , DNA Helicases , DNA Bacteriano/genética , Proteínas de Ligação a DNA , Escherichia coli/genética , Plasmídeos , Pseudomonas aeruginosa/genética , Transativadores , Sequência de Aminoácidos , Proteínas de Bactérias/metabolismo , Sequência de Bases , Sítios de Ligação , Calorimetria , Clonagem Molecular , Replicação do DNA/genética , DNA Bacteriano/química , Vetores Genéticos , Genótipo , Dados de Sequência Molecular , Mutagênese Insercional , Oligodesoxirribonucleotídeos , Mapeamento por Restrição , Deleção de Sequência , Homologia de Sequência de AminoácidosRESUMO
Sequence analysis of the Pseudomonas aeruginosa insertion sequence element IS222 revealed it to be 1234 bp in size with 23 bp imperfect terminal inverted repeats. Insertion caused a 5-bp duplication of the insertion site. Two ORFs were identified, one of which, ORFA, could encode a basic (pI 10.5) polypeptide with a mass of 11,709. This sequence bears strong homology to the putative ORFA product from the Shigella dysenteriae insertion sequence element IS911, which is a member of the IS3 family of insertion elements. As with other members of this group the nucleotide sequence contains a "frameshift window" (AAAAAAG; M. Chandler and O. Fayet (1993). Mol. Microbiol. 7, 497-503) at which ribosome slippage can result in a fusion protein (ORFAB).
